Silver Nano in Biomedical

CHAPTER 3
Nanoparticles for Biomedical Applications

Tianzhong Yang, Anca Mocofanescu, Chengmin Shen, Hongjun Gao, and
Congwen Xiao
3.1
Introduction
Particles with diameters in the range of 2 to 100 nm, so-called nanocrystalline materials, have become a major interdisciplinary area of research during recent decades.
In fact, since the seventeenth century, noble metallic nanomaterials, though not
understood, have been obtained and used to give rise to a brilliant rose color
throughout Europe in stained glass windows of cathedrals and by the Chinese in
coloring vases and other ornaments [1, 2]. The scientific preparation of nanoparticles dates back to the nineteenth century, with Faraday reporting the preparation of colloids of relatively monodispersed gold nanoparticles. The scientists who
major in nanoscience and nanotechnology should appreciate the inventors who
designed transmission electron microscopy (TEM). The high-resolution TEM
(HRTEM) and low-resolution (LRTEM) allow one to observe a substance at a
nanometer scale directly. The magic machines make it possible to investigate the
nanomaterials with respect to size, size distribution, shape evolution, and shape uniformity and even the structure. The past couple of decades have witnessed an exponential growth of activities in this field worldwide, driven both by the excitement of
understanding new science and by the potential hope for applications and economic
impacts. Indeed, many efforts have been devoted to investigation into the synthesis,
characterization, and application of nanomaterials. In general, nanomaterials can
be classified into three groups: zero-dimensional materials, so-called nanoparticles,
with variations in shape and diameter, one-dimensional materials, including
nanorod and nanowire, and two-dimensional materials, including nanobelts,
nanodisks, films, and nanosheets. Herein we focus on the nanoparticles (NPs), especially the metallic ones.
The intense interest in the metallic NPs derives from their unique chemical and
electronic properties arising from the small volume to big surface area ratio and the
separation in the electronic energy level. The change in the properties at this length
scale from their bulk counterparts is not a result of only scaling factors. It results
from different causes as far as different materials are concerned. In semiconductors,
it results from the further confinement of the electronic motion to a length scale that
is comparable to or smaller than the length scale characterizing the electronic
motion in bulk semiconducting material (called the electron Bohr radius, which is
43
44
usually a few nanometers). As noble metals are reduced in size to tens of

nanometers, a new very strong absorption is observed, resulting from the collective
oscillation of the electrons in the conduction band from one surface of the particle to
the other. This oscillation has a frequency that absorbs the visible light. This is called
the surface plasmon absorption. In the case of transition metal nanoparticles, the
decrease in the particle size to the nanometer length scale increases the surfaceto-volume ratio. This, together with our ability to make them in different sizes and
shapes, makes them potentially useful in the field of catalysis.
The absorption spectra of many metallic nanoparticles are characterized by a
strong broad absorption band that is absent in the bulk spectra. Classically, this
giant dipole (or surface plasmon) band is ascribed to a collective oscillation of the
conduction electrons in response to optical excitation [3]. The presence of this band
in the visible region of the spectrum is responsible for the striking colors of dilute
colloidal solutions of noble, alkali, alkaline Earth (Ca, Sr, Ba), and rare Earth (Eu,
Yb) nanoparticles [4]. Mies theory predicts that below a certain size, less than
one-tenth of the optical wavelength, the position and width of this band should
remain constant, independent of size [5]. Experimental evidence, however, together
with a dramatic increase in width with decreasing size [6], indicates a slight but significant shift to lower energy. Fragstein and Kreibig and others [79] have investigated the optical absorption spectra theoretically in the case of free-electron metals
and advanced the theory that when the particle diameter becomes small enough,
with diameter less than the electronic mean-free path in the bulk metal (~20 nm for
gold), the scattering of free electrons with the particle surface begins to have an
influence on optical excitation. Such a simple and practical theory succeeds in terms
of spectra of relatively large particles (>3 nm for gold) but does not agree with the
experimental results when applied to the smaller sizes. It is anticipated that at one
point the phenomenological description of free electrons, as well as inherent fundamental assumptions of infinite lattice periodicity and a continuous energy level spectrum, must fail, because it is widely known that the continuous energy level band
will separate into individual levels since in the cluster the number of atoms is so
small. Experimental measurements for the small nanoparticles are often degraded
by a lack of size and shape uniformity that renders comparison with theory questionable [10], and this is what makes it so difficult to unequivocally identify quantum size effects in the optical spectra of metal nanoparticles prepared in
macroscopic quantities, although such effects are well known from experiments on
metal-cluster beams and from conductance measurements on single-metal
nanostructures [11].
Hence, it is a prerequisite to take a good grasp on the control of the size distribution of the metallic nanoparticle. In later discussion, we see the same problem in the
case of magnetic metallic nanoparticles. So much effort has been devoted to finding
an approach that can afford us nanoparticles with a narrow size distribution.
Besides noble metal nanoparticles, nanoparticles with novel magnetic properties
are attracting more and more attention with regard to bioapplications. As far as
magnetism is concerned, magnetic particles with diameters smaller than some certain critical value usually show properties different from their bulk counterparts. At
room temperature, the measured M-H curve, namely the hysteresis loop, recorded
on the magnetic nanoparticles shows saturation magnetization but no coercive field
3.2 Synthesis of Metallic Nanoparticles
45
or magnetic remanence synchronously. In other words, from the viewpoint of the

coercive field or magnetic remanence, they behave similarly to paramagnetic materials; on the other hand, the magnetization can reach the saturation state and is
larger than paramagnetic ones. The scientists describe this behavior as
superparamagnetic. This indicates that magnetic moments in the particles are free
to align with the field during the measuring time at room temperature. This phenomenon is caused by the fact that in such small particles, because of the thermal
fluctuation the magnetic moments can rotate freely despite the magnetic energy barrier. This characteristic allows a promising future in application of the magnetic
nanoparticles in biomedicine, particularly in magnetic resonance imaging (MRI),
tissue engineering, and drug delivery.
Because of both the excitement of understanding a new science and its potential
for applications having economic impact, a great deal of effort has been devoted to
investigations into the metallic nanoparticles in terms of synthesis, size and shape
control, surface modification, and the primary applications. Significant breakthroughs have been achieved in the synthesis of nanoparticles of varying diameters
and in obtaining a narrow size distribution. Based on these, highly ordered
self-assembly structures with large area of metallic nanoparticles have been
obtained. For the purpose of bioapplication, some kinds of organic molecules,
including biomolecules of different lengths and functional groups, have been coated
or linked onto the surface of metallic nanoparticles.
In this review, we discuss the synthesis and properties of zero-dimensional
metallic nanomaterials, including the noble metal nanoparticles and the magnetic
ones. We start with a discussion of some effective methods aimed at the synthesis of
the nanoparticles. After that their novel properties are introduced. The application
of the metal nanoparticles in biotechnology is presented in the final section. We
focus on the synthesis of metallic nanoparticles by the wet chemistry method.
Chemical vapor deposition (CVD) and the physical techniques such as using electron, ion, or photon beams in lithography to make nanostructures are not discussed.
In general, this chapter is constructed based on the remarkable work reported in the
past decades. Because of the limitation of our knowledge and experiences, there
must be some incorrect sayings or points in this overview. Due to the explosion of
publications in this field, we do not claim that this review includes all of the published work, but rather an exposure to the methods. We apologize to other authors
who have contributed to the synthesis, characterization, and application of the
metal NPs whom we have unintentionally left out.
3.2
Synthesis of Metallic Nanoparticles

3.2.1
Synthesis Approaches to Noble Metal Nanoparticles
In fact, as mentioned earlier, since the seventeenth century, noble metallic

nanomaterials have been observed and used, but not understood, to give rise to a
brilliant rose color throughout Europe in stained glass windows of cathedrals and
by the Chinese in coloring vases and other ornaments. And the scientific preparation of nanoparticles dates back to the nineteenth century, with Faraday reporting
the preparation of colloids of relatively monodispersed gold nanoparticles. In the
46
past decades, many approaches have been utilized to produce the noble nanoparticles. Before moving into the idiographic synthesis methods, we discuss the particles growth in kinetics and the mechanism.
3.2.1.1
Introduction
In general, the process of generation of a solid phase from solution inevitably

involves in the chemical growth nanometer-sized materials. Two primary steps constitute the course of NPs growth: Step 1, the nucleation occur; step 2, by adsorbing
the substance separated out of the solution, the nucleoli grow into the particles of
desired size and shape. A good understanding of the process and parameters demonstrating the precipitation helps to tailor the growth of nanoparticles to the desired
size and shape. For a particular solvent, there is certain solubility for a solute,
whereby addition of any excess solute will result in precipitation and formation of
nanocrystals. Thus, in the case of nanoparticle formation, for nucleation to occur,
the solution must be supersaturated either by directly dissolving the solute at higher
temperature and then cooling to low temperatures or by adding the necessary reactants to produce a supersaturated solution during the reaction [12, 13]. The precipitation process then basically consists of a nucleation step followed by particle
growth stages [14, 15].
Generally, there are three kinds of nucleation processes: homogeneous, heterogeneous, and secondary. Homogeneous nucleation takes place in the absence of a
solid interface by combining solute molecules to produce nuclei, and this procedure
is involved in the synthesis of metallic NPs by the wet chemistry method. Homogeneous nucleation happens due to the driving force of the thermodynamics because
the supersaturated solution is not stable in energy. The overall free energy change,
G, which dominates the nucleation progress, is the sum of the free energy due to
the formation of a new volume and the free energy due to the new surface created.
After the nuclei are formed from the solution, they grow via molecular addition,
which relives the supersaturated step. On the other hand, when the reactants are
depleted due to particle growth, Ostwald ripening or defocusing will occur, where
the larger particles continue to grow and the smaller ones get smaller and finally dissolve. In addition to the growth by molecular addition, where soluble species deposit
on the solid surface, particles can grow by aggregating with other particles, and this
is called secondary growth. The rate of particle growth by aggregation is much
larger than that by molecular addition. After the particles grow to a stable size, they
will grow by combining with smaller unstable nuclei and not by collisions with other
stable particles through such a mechanism of oriented attachment of nanocrystals
[16, 17].
3.2.1.2
Synthesis of Gold Nanoparticles
The synthesis of colloidal gold has been studied intensively for a long time [18]. In
1994, Brust et al. [19] developed an effective method for the generation of
long-chain alkanethiolate-stabilized gold nanoparticles. The product from this system is easy to disperse in organic solvent and to reisolate as pure powders. Due to the
superb stability of the as-prepared particles, many groups have tried to follow or
47
develop the approach. Much work has been done based on this smart design to
modify the properties, such as the reactivity and solubility, of the nanoparticles
through changing the molecular structures of the thiolates on the particle surface
[2026]. Both long-chain thiolates containing an aromatic moiety [20], -substituted (cyano, bromo, vinyl, ferrocenyl) [21] and poly-hetero--functionalized [22]
long-chain alkanethiolates and small molecular thiolates such as ( -mercaptopropyl) trimethyloxysilane [23] and the rigid aromatic thiols of 4-mercaptobiphenyl [24] and p-mercaptophenol [25] have been utilized. Kunitake and
coworkers also showed that sodium 3-mercaptopropionate could be used for the
synthesis gold nanoparticles using citrate as reductant [26]. Since the diluted aqueous solution was needed in this method, it is difficult to separate the particles as
solid powders; therefore, the detailed characterization of the surface structure with
a variety of methods where solid powders are needed as the specimens is prevented.
In addition, regarding the application of the gold nanoparticles in biomedicine,
where the pH value is close to neutral, there still will be the challenge of developing
the approach from which the Au nanoparticles can be easily dispersed into aqueous
solution at a neutral pH value.
In 1999, Sihai Chen and Keisaku Kimura reported a new approach that can
avoid the disadvantages mentioned above [27]. The shining points of their work are
the standout elucidation of the surface structure of these particles and that the
as-synthesized nanoparticles are easily dispersible in water, a property that has not
been achieved using other kinds of methods where the thiolates are employed
[2026]. For biomedical purposes, the latter feature is a prerequisite because the
nanoparticles should combine with macromolecules in living or fixed cells or tissues
in aqueous solution.
Let us introduce the process briefly. In a typical preparation, aqueous solution
was at first mixed with mercaptosuccinic acid (MSA) in methanol to give a transparent solution. Under vigorous stirring a freshly prepared aqueous sodium
borohydride solution was then added at a certain rate. The solution turned dark
brown immediately but remained transparent (which indicates that the nanoparticles do not form) until enough reductant was added. The pH of the solution
increased gradually with the addition of reactant. A flocculent dark brown precipitate would be generated by further addition of the reductant, and finally the pH of
the solution was brought to 8.6. We can see that just as in the common synthesis in
aqueous solution, the pH must be adjusted. After being stirred for another duration,
the solvent was removed by decantation after the centrifugation. The precipitate
was washed, suspended in ethanol, and dried by rotary evaporation without
exceeding a temperature of 40C under vacuum and at last the product in powder
state was obtained.
They explained the whole procedure as follows:
8nHAuCL 4 + 8mR SH + 3nNaBH 4 + 12nH 2 O
8( Au ) n R S
+ 3nB(OH) 4 + (32n + 8m)H + + 32nCl + 3nNa +
(3.1)
The initial molar ratio between MSA and chloroauric acid plays a dominate role in
tailoring the relative rates of particle nucleation and (then) growth, and hence it
finally determined the particle size. It was found that the particle sizes decreased
48
with the increase in S/Au ratios. This result is similar to that reported by Leff et al. on
the long-chain alkylthiol-passivated gold nanoparticles [28], suggesting that the
thermodynamics are responsible for the particle stability.
One can see that H+ ions were released in reaction 1, while the final pH of the
reaction mixture increased with the addition of NaBH4 due to the strong hydrolysis
of NaBH4, which can be elucidated as follows:
NaBH 4 + 4H 2 O NaOH + 4H 2
(3.2)
Hence, the final pH is determined by the competition between reactions (3.1) and
(3.2). We keep our eyes on the pH value because it influences the dispersibility of the
gold nanoparticles, which is important in terms of biomedical applications. For
instance, the dispersibility of gold particles abruptly decreased when the pH of the
solution increased to around 5 during the addition of NaBH4. Furthermore, due to
these pH-dependent properties, gold particles can be easily separated from inorganic
and organic impurities by increasing the pH of the solution.
The notable property of the as-synthesized nanoparticles is that they are easily
dispersed in neutral water, since before this report gold nanoparticles were found to
be dispersible only in alkaline aqueous solution (pH 13) using other kinds of
thiolate such as p-mercaptophenol [29]. For the bioapplication purposes being considered, the pH value is too high. However, the size-dependent dispersibility of the
powder in water (that is, its increase as the particle size decreases) may be a shortcoming rather than an advantage because it suggests a poor MSA coating on the surfaces of larger gold nanoparticles, which means that the larger particles from this
method may be not suitable for bioapplications.
As mentioned in the introduction section, one problem is that experimental
measurements for the small nanoparticles are often degraded by a lack of size and
shape uniformity that renders comparison with theory questionable [30]. Hence, it
is almost impossible to unequivocally identify quantum size effects in the optical
spectra of metal nanoparticles prepared in macroscopic quantities, although such
effects are well known from experiments on metal-cluster beams and from conductance measurements on single-metal nanostructures [31].
As the saying goes, where there is a will, there is a way. The discovery of methods for preparing gold nanoparticles by Dubois and Nuzzo in 1992 [32] has overcome this obstacle successfully. Gold alkylthiolate assemblies that are a
nanometer-scale analog to the well-studied surface system were utilized. One notable point in this synthesis is that on the extended Au surfaces, one finds an extraordinary example of the uniform protection of the surface without modification of its
essential structural and electronic properties. N-Alkylthiolates and their derivatives
form compact, ordered monolayers where thiolates (RS-) or dialkyl disulfides
(RSSR) reversibly attach to various Au surfaces. Because of favorable (van der
Waals) interactions among the long, ordered alkane chains in concert with a weak
chemisorption of the sulfur head group to the metal surface (~1.2 eV per RSSR unit
desorbed versus ~10 eV surface energy per six Au atoms covered by this unit), the
highly ordered self-assembly occurs spontaneously. These extensively studied systems have recently begun to be well understood. For instance, a surface structure
49
implicating interactions between Au surface atoms and the nonbonding S orbitals of

an intact RSSR molecule was identified by Fenter et al. in 1994 [33].
In 2001, a report from Hongjun Gaos group presented outstanding work on
the Ag nanoparticles with narrow size distribution. And based on this characterization, the self-organized superstructure was obtained. From Figure 3.1 one can see
that the NPs have a diameter of about 4.18 nm and the standard deviation of the
size distribution is 0.23 [34].
3.2.2
Synthesis of Magnetic Metal Nanoparticles
Population (%)
Magnetic nanoparticles, especially those with diameters of less than 10 nm, are of
intense interest because the particle size is smaller than the characteristic dimensions
such as atomic or ionic diffusion length, electronic elastic and inelastic mean free
path length, and the magnetic domain size [3539]. Many new phenomena and
properties are expected to appear in the NP systems. The size and shape specificity
of nanoparticles naturally acts as building blocks of the self-assembled passivated
NP superlattices (SLs) or NP arrays [40]. In this emerging field, monodisperse magnetic NPs are a prerequisite to a broad range of applications, such as data storage
devices, biosensors, and drug delivery [4144]. The advances in preparing semiconductor and metallic NPs, specifically by the method of injecting molecular precursors into hot organic surfactant solution, have improved the NP samples
remarkably, with good size control, narrow size distribution, and good crystallinity
of disperse nanoparticles [45, 46].
Now we introduce the general approach though the synthesis of cobalt
nanoparticles. The reason we chose to begin with the Co system is that Co nanocrystals possess a wealth of size-dependent structural, magnetic, electronic, and
30
25
20
15
10
5
0
90 nm
3.0 3.5 4.0 4.5 5.0 5.5
Diameter (nm)
Figure 3.1 TEM image of a two-dimensional silver nanoparticle superlattice and (inset) the histogram of the nanoparticles. (Source: [34].)
50
catalytic properties. In particular, the exponential dependence of the magnetization

relaxation time on volume has motivated intensive studies of Co nanocrystal synthesis for magnetic storage purposes. It is difficult to make isolated magnetic nanocrystals of Co, in part because the forces between/among the particles are large.
These forces occur due to both the high electron affinity and the high surface tension
arising from the partially filled D-band, the large van der Waals forces between
polarizable metal particles, and magnetic dipole interactions [45].
In addition, a new structure has been found for cobalt at the nanometer scale. It
has been well known that cobalt (bulk) has two crystal structures, close-packed hexagonal (hcp) and face-centered cubic (fcc), that both phases can coexist at room temperature, and that the fcc structure is thermodynamically preferred above 450C
while the hcp phase is favored at lower temperatures. In the case of small particles,
however, the fcc structure appears to be preferred even below room temperature
[47]. The existence of fcc and hcp cobalt was first reported by Hull [48] in 1921 by
analyzing diffraction patterns of metallic powders obtained by several methods.
Half a century later, in 1974, Krainer and Robitsch reported the observation of new
diffraction lines in samples prepared by spark erosion of bulk cobalt surfaces; unfortunately the structure was never fully established [49]. After another decade passed,
Kajiwara et al. [50] noted several new lines in the diffraction pattern of cobalt
nanoclusters prepared by plasma evaporation and the subsequent condensation of
the metal; they attributed these lines to a form polymorphous to the two known
structures. Similar results are reported by Leslie-Pelecky et al. [51] with cobalt particles prepared by the reduction of cobalt salt in solution with metallic lithium. It is
from 1976 that the evidence for a new structure of cobalt present in small cobalt
clusters produced by the decomposition of organometallic precursors is provided
by Respaud et al. [52]; however, the structure was not identified. In 1999, Dmitry
P. Dinega and M. G. Bawendi reported and identified a new stable structure for
elemental cobalt [53]. Now it is well known as -cobalt. A detailed discussion of
this standout work will be given later; at this stage, we want to talk about the
relatively general synthesis of nanoparticles, especially by means of so-called wet
chemistry.
It is well known that inert atmosphere solution phase metal salt reduction is a
powerful technique for the preparation metal particles. For controlling particle
growth, stabilizing metal dispersions, and limiting oxidation of the particles, many
ligands, including polymers and surfactants, have been employed. It is obligatory for
the stability of the colloid particles that repulsive forces of sufficient strength and
range exist to counteract the combined attractive forces. In the case of metal particles, the strong van der Waals attractions that combine with the magnetic dipole
interactions make it challenging to stabilize these systems. To overcome this, many
approaches have been proposed and utilized. For example, the employment of a
combination of surfactantstrialkylphosphine and oleic acidwas realized to control particle growth, stabilize the particles, and prevent oxidation [54].
When the experimental conditions and the organic coatings are adjusted to be
suitable, nanoparticles of uniform size, shape, composition, and structure can be
obtained, which are necessary for both fundamental and application aims. When
these are achieved, it is possible to obtain the superstructure of these particles, so
called self-assembly, in a one-, two-, or three-dimensional manner. The
51
superstructures possess lots of advantages in application in bionanotechnology and

the recording industry.
For the purpose of inducing three-dimensional assemblies [55, 56], controlling
the deposition of the nanoparticles on various substrates, ensuring their biocompatibility, and forming an airtight shell that would protect the NPs from oxidation,
many scientific groups focus on tailoring nanoparticle surface. Chemical control of
the synthesis of nanometer scale materials also results in fascinating phenomena
such as stabilization of new phases [57] and change in electronic and magnetic
properties due to finite size and strong surface effects related to the coordination of
ligands at the NP surface [58, 59]. These considerations are especially important
when we take into account that the anisotropy and magnetization of the magnetic
NPs depend on the structure and the volume. Furthermore, anisotropic growth of
the NPs is claimed to be controlled via ligand coordination in certain cases [60]. It is
thus crucial to obtain a better knowledge of the surface/ligand interaction. Hence, it
is necessary to become acquainted with the synthesis methods and the corresponding modifications before moving into the properties, which are dependent on the
structure, the dimension, the modification of surface, and the bioapplication
sequentially. Let us begin by introducing Bruno Chaudret and coworkers work.
They have long been involved in the synthesis of metal NPs employing an
organometallic route and more specifically in the study of ligand coordination at
the surface of the NPs [61, 62], together with the effect of such a coordination on
their structure [63, 64] and magnetic properties [65, 66]. In 1996, they changed the
structure of Pt NPs from fcc to icosahedral with the utilization of the phosphine coverage on their surfaces [64]. In the same year, they found the collapse of cobalt NP
magnetization, which was induced by adsorption of carbon monoxide at the surface
of the NPs embedded in polymer matrices [65].
As far as the limitation that only very small molecules are able to diffuse
through the polymer matrix is concerned, employing ligands as stabilizing agents
during the NP synthesis thus came into their view for the purpose of further investigations of the relationship between the chemical nature of the ligands and their
influence on surface magnetism. They showed that donor ligands such as amines did
not alter the surface magnetism of the NPs while promoting the formation of
nanorods as a result of shape control, whereas the use of trioctylphosphine oxide
led to NPs of reduced magnetization, in the case of nickel NPs in 2001 [67].
In 2005, they reported the synthesis, structure, and magnetic properties of
cobalt NPs (sample 1) obtained from two different precursors, Co(3C8H13)
(4C8H12) and Co[N(SiMe3)2]2, in the presence of di-isobutyl aluminum hydride
(DiBAH), a commercially available aluminum-based Lewis acid, and the results
were compared with those obtained when the NPs were synthesized in the commonly used polymer matrix poly-(vinylpyrrolidone) (PVP) [65, 66]. The choice of
an aluminum compound here is based on the fact that the Al compound can be the
starting point for the growth of alumina coatings, which should display interesting
dielectric properties and high thermal conductivity and possibly prevent air oxidation [68].
The observed difference in the structures can be explained based on consideration of the different binding behaviors of the organic coating. NPs with a structure
similar to that of the product from the precursor of Co(3C8H13)(4C8H12) in the
52
presence of DiBAH have been reported by them [69]. In that work, they obtained Co
nanoparticles with small mean diameters of about 1.4 nm, by means of hydrogenation of Co(3C8H13)(4C8H12) companied by poly(vinyl pyrrolidone). A comprehensive study of their magnetic properties has shown that they are similar to those
expected for free-standing cobalt NPs, which were discussed in 1998 [70]. Briefly, if
one assumed that the polymer matrix has few or no interactions with the NPs, the
deduction is that surface metal atoms are undercoordinated and electronically
unsaturated. The noteworthy point is that the formation of -cobalt has been
reported to be promoted by a careful choice of the ligands used during the synthesis
[53, 54], which will be discussed latter. Hence, in the presence of trioctylphosphane
oxide (TOPO) or PR3 ligands, which are known to damp the magnetic properties of
NPs, cobalt NPs adopt the -cobalt structure, whereas an fcc structure is observed in
the absence of such ligands [70]. In sample 1, the electron deficiency is also important. On the contrary, NPs in sample 2 (when [CoN(SiMe3)2]2 is utilized as precursor; for detail, see [71]), the surfaces of which are surrounded by donating ligands,
adopt a compact structure displaying features of both fcc and hcp phases. It is worth
noting that the hcp structure is not often reported for such small sizes, especially for
NPs obtained at low temperature. It is suggested that coordination of a -donor
ligand at their surfaces favors this structure. The synthesis conditions also play an
important role on the structure of the product. In fact, sample 1 is obtained at room
temperature by a slow reaction process (48 hours), whereas the formation of sample
2 is fast at 50C. This is in contradiction with the expected kinetic quenching of a
metastable structure and further attests the influence of the coordinated ligand on
the structure adopted by the NPs.
Now let us come to the identification of the new phase of cobalt in detail. As was
mentioned, besides the well known structures including hcp and fcc, another phase
of cobalt has been found in cobalt nanomaterials. Though the insight was reported
by Hull [72] in 1921 and then Krainer [73] and Kajiwara et al. [50], it is from 1976
that the evidence for a new structure of cobalt present in small cobalt clusters produced by the decomposition of organometallic precursors is provided by Respaud et
al. [74]. However, the structure was not identified until Dinega and Bawendi
reported their work on this new structural cobalt in 1999 [53]. In the report, thermal
decomposition of octacarbonyldicobalt in solution in the presence of TOPO as a
coordinating ligand was employed to synthesize cobalt nanoparticles. This method
provides a clean route for the preparation of the material since elemental cobalt is
the only nonvolatile product of the reaction, which can be depicted as follows:
[Co
(CO) 8 ] 2Co + 18CO
(3.3)
They claimed that the as-synthesized powder was highly susceptible to magnetic
fields generated by a small permanent magnet, suggesting that it consisted of magnetic materials without or with thimbleful of cobalt oxides. TEM showed that the
powder consisted of roughly spherical particles with an average diameter of 20 nm
and a 15% size distribution. Later experiments showed that the size of the crystals is
not limited to this range. In fact, injection at higher temperature yields regular
polyhedron-shaped faceted crystals as large as 0.3 m and possibly even larger
(Figure 3.3). These results are reasonable since we know that the temperature will
611
442
610
531
510
511
330
420
321
210
211
110
520
521
310
53
311
221
(a)
(b)
(c)
20
30
40
50
60
70
80
90
100
2/
Figure 3.2 (a) Experimental and (b) calculated X-ray powder diffraction patterns of the -cobalt
structure and (c) the sample shown in (a) after being heated to 500C. Peaks corresponding to fcc
cobalt are denoted by X. (From: [53]. 1999 Wiley-VCH Verlag GmbH, Weinheim, Germany.
Reprinted with permission.)
influence the nucleation and the growth process, as discussed previously. They
also found the presence of oxygen, which indicates surface oxidation of the cobalt
particles, corresponding to coverage of one to two monolayers of cobalt oxide for
particle sizes in the 20-nm range. The other elements, in their opinion, come from
residual organic solvents adsorbed on the particle surfaces. The absence of phosphorus in the elemental analysis indicates that TOPO was completely removed during washing of the particles. It is noted in this work that freshly synthesized cobalt
nanoparticles are extremely reactive toward oxidation. In fact, rapid exposure to air
results in immediate oxidation accompanied by a red glow. Therefore, even simply
washing the particles in organic solvents inevitably leads to surface oxidation by
residual moisture and dissolved air. The consideration is reasonable that the resulting oxide layer appears to passivate the surface of the particles and considerably
reduce the speed of further oxidation. It should be mentioned that the new -cobalt
phase appears to be metastable under normal conditions. Although stable at room
temperature for at least several months, heating the sample at 500C completely
transforms it to the known fcc phase. Furthermore, subsequent cooling does not
return the sample to its original -cobalt structure.
Figure 3.2 shows the X-ray powder diffraction patterns of the as-synthesized
product, which present the evolution of the nanoparticle structure with variation of
the molar ratio of TOPO/precursor, and one can see that this product takes a new
54
phase that does not correspond to either of the two known structures of cobalt, fcc
and hcp. In other words, neither the positions of the peaks nor their relative intensities correspond to any known cobalt phases. Based on the detailed analysis, the conclusion is that this pattern corresponds to a new structure of cobalt, so-called
-cobalt. This structure is cubic (space group P4132) with a unit cell parameter =
6.097 0.001 . The unit cell structure is similar to that of -manganese (a
high-temperature phase of manganese) [75]. Twenty cobalt atoms constitute one
unit cell, and they are divided into two types: 12 atoms of type I and 8 atoms of type
II. These two types of atoms differ in their local coordination. Unlike an ideal
close-packed structure, which has 12 nearest neighbors, -cobalt has only three
nearest neighbors for type I and two nearest neighbors for type II atoms. This phenomenonthe same atoms being divided into several types based on the local coordinationoften happens in compounds such as magnetite. This results in a structure
for -cobalt that is less dense than both the hcp and fcc structures. The calculated
density of -cobalt is 8.635 g(cm)3, compared to 8.788 g(cm)3 and 8.836 g(cm)3
for the fcc and hcp structures, respectively [53].
Hence, the structure of -cobalt was identified for a certainty. Figure 3.3 shows
the TEM images of the as-obtained nanoparticles. However, as far as the uniformity
of the size and shape is concerned, which we continue to emphasize, the system
needs to be developed for the uniformity of the morphology.
Figure 3.4 shows models of the unit cell for -cobalt with respect to different lattice direction projections. In addition, for the purpose of applications, it should be
mentioned that the magnetic moment per atom in -cobalt, measured as 1.70 B, is
similar, within experimental error, to those of the two known structures (1.75 and
1.72 B for bulk fcc and hcp cobalt) [53, 76].
It was also claimed that the use of TOPO seized the key role here because of the
fact that the same preparation method but without the presence of TOPO yields
nanocrystals with a pure fcc phase. Hence, the influence of TOPO on the structure
of the as-obtained nanoparticles should be clarified, and it was indeed done. Tight
coordination of ligand molecules (TOPO) around the growing crystal and around
solubilized cobalt atoms is responsible for changing the energetics of growth in favor
of the new, less dense phase. However, the particular synthesis of this paper is not
unique in producing -cobalt. Sun and Murray [54] have confirmed this assignment
0.1
0.1m
Figure 3.3 TEM image of cobalt crystals with the new -cobalt structure. (From: [53]. 1999
Wiley-VCH Verlag GmbH, Weinheim, Germany. Reprinted with permission.)
(a)
55
(b)
Figure 3.4 The unit cell of -cobalt: (a) unit cell cube filled with 8 atoms of type I (dark) and 12
atoms of type II (light) and (b) 111 projection of the same cube showing threefold symmetry along
its main diagonal. (From: [53]. 1999 Wiley-VCH Verlag GmbH, Weinheim, Germany. Reprinted
with permission.)
of the structure of -cobalt in nanocrystals obtained by reducing cobalt salts in solution in the presence of alkylphosphanes serving as the coordinating ligands. Due to
its metastability, -cobalt may be accessible only by solution-phase approaches,
rather than more common techniques, such as electrochemical deposition or chemical vapor deposition and so on. As far as the uniformity of nanoparticle morphology is concerned, remarkable achievements have been obtained by Sun and
Murrays work. In their work, briefly, the particle generation began with the injection of dioctylether superhydride solution into a hot (200C) CoCl2 dioctylether
solution in the presence of oleic acid (octadec-9-ene-1-carboxylic acid,
CH3(CH2)7CH=CH(CH2)7COOH) and trialkylphosphine (PR3, R=n-C4H9, or
n-C8H17). Reduction occurred instantly upon injection, leading to the simultaneous
formation of many small metal clusters serving as nuclei. Continued heating at
200C allowed steady growth of these clusters into nanometer-sized, single crystals
of cobalt. The steric bulk of the alkylphosphine controlled the rate of particle
growth and consequently the morphology. Short-chain alkylphosphines (e.g.,
tributylphosphine) allowed faster growth, leading to the generation of large particles, while bulkier species (e.g., trioctylphosphine) reduced particle growth and
favored production of smaller nanocrystals (NCs). However, it is worth mentioning
that the influence of the organic coatings steric hindrance on the particles size, size
distribution, and shape must play some essential role in such cases. These organically stabilized cobalt NCs were readily dispersed in aliphatic, aromatic, and chlorinated solvents and could be precipitated with the addition of short-chain alcohols,
facilitating a size-sorting procedure that isolated nearly monodisperse samples [77].
In Sun and Murrays work, average particle size was coarsely controlled by
varying the type of alkylphosphine used in combination with oleic acid during the
growth. For instance, bulky P(C8H17)3 limited growth to producing particles with
diameters of 2 to 6 nm, and less bulky P(C4H9)3 led to larger particles (7 to 11 nm in
diameter). A size selection procedure is needed for the purpose of fine tuning the
particle size. It should be pointed out that the size sorting procedure is the disadvantage of such a system in view of the cost for application in industry. And some
groups developed approaches that lead to nanoparticles with tunable size and narrow size distribution without the size selection procedure. Based on the fact that
trialkylphosphine reversibly coordinates neutral metal surface sites, slowing but not
56
stopping the particles growth, it cannot prevent the particles from eventually growing to undispersible aggregates at high temperature when used alone. If oleic acid is
employed alone, though it is an excellent stabilizing agent, it binds so tightly to the
particle surface during synthesis that it greatly impedes the particle growth. The
combination of trialkylphosphine and oleic acid produced a tight ligand shell that
allowed the particles to grow steadily while it protected them from aggregation and
oxidation.
It is well known that a temporal separation of the nucleation and growth stages
is needed to produce a monodisperse colloid. Ideally, a large number of critical
nuclei should be formed in a short interval of time followed by the simultaneous and
steady growth of those nuclei. In Sun and Murrays approach [54], the reducing
agent (superhydride) was injected to induce rapid reduction of cobalt (II) chloride in
hot (200C) media. This procedure provided the temporally discrete homogeneous
nucleation desired. Growth of the nuclei was continued by addition of cobalt-containing species to the surface of the particles and was halted by cooling to below
100C naturally. From the TEM characterization (Figure 3.5), one can see the standout results obtained by this means. However, the size selection procedure is needed.
This may lead to a high cost in terms of economics when it is realized in industry.
In 2003, a report from Gaos group presented the advancement in the control of
size, size distribution, and the monolayer and multilayer self-assembly based on the
narrow size distribution [78, 79]. In their work, they employed the approach in
which thermal decomposition of octacarbonyldicobalt was utilized. This method
provides a clean route for the preparation of the material since the cobalt is the
only nonvolatile product during the whole and the reaction can be described by (3.3)
[53, 80]:
A combination of surfactants in the presence of stabilizing ligands triphenylphosphine and oleic acid was employed for controlling the particle growth, stabilizing the particles, and preventing oxidation. Because the phenyl can provide greater
steric hindrance than that of the straight-chain alkyl, they adopted the triphenylphosphine, instead of the tributylphosphine or trioctylphosphine, to synthesize
cobalt NPs with smaller size.
48nm
Figure 3.5 TEM image of a two-dimensional assembly of 9-nm cobalt nanocrystals. Inset:
High-resolution TEM image of a single particle. (From: [54]. 1999 American Institute of Physics.
3.3 Novel Properties of Metal Nanoparticles
57
Briefly, a three-neck flask with dichlorobenzene is heated to ~220C under N2

and stirred for a certain time to exclude ambient air, and then equivalent amounts in
mole of oleic acid and triphenylphosphine are added. In another vessel, Co2(CO)8 is
combined with dry dichlorobenzene, warmed until fully dissolved, and then afterward rapidly injected into the flask. The solution turns black quickly and foams,
indicating the precursor decomposition and the nucleation. The black solution is
refluxed and then the particles are obtained by the classical separation procedure.
Cobalt nanoparticles with 7-nm average diameter are obtained by the size-sorting
procedure with a narrow size distribution. When the concentration of the oleic acid
and triphenylphosphine was increased by a factor of 2, cobalt NPs of about 5 nm
were obtained.
Compared to Dinega and Bawendis work [53], at first the employment of oleic
acid played a role in leading to the differences. As we have discussed, oleic acid,
when employed alone, is an excellent stabilizing agent. However, it binds so tightly
to the particle surface during synthesis that it greatly impedes the particle growth.
The combination of triphenylphosphine and oleic acid produced a tight ligand shell
that allowed the particles to grow steadily while protecting them from aggregation
and oxidation. Second, by using the triphenylphosphine instead of the tributylphosphine or trioctylphosphine, based on the fact that the phenyl can provide
greater steric hindrance than that of the straight-chain alkyl, the nanoparticles with
smaller diameter (7 nm) were obtained. In addition, since it is generally accepted
that a temporal separation of the nucleation and the growth stages is required for
the production of a monodisperse colloid of nanoparticles [54], the injection of the
dichlorobenzene solution of Co2(Co)8 into a hot system (~220C) forms a large
number of critical nuclei. The growth of the nuclei continues by the addition of
cobalt-containing species to the surface of the particles with a decreasing reaction
rate (temperature tuned to 185C). This process can narrow the size distribution of
cobalt NP diameters. Figure 3.6 exhibits the highly ordered superlattice structure of
the cobalt nanoparticles.
3.3
Novel Properties of Metal Nanoparticles

As we all know, the composition and the structure determine the properties, and the
properties determine where and how the materials can be used. So it is necessary to
know about the properties before moving into the applications.
3.3.1
Unique Properties of Noble Metal Nanoparticles
In the following we will locate space to discuss the properties of metal nanoparticles, especially those with organic coating, because these underlie the application of the nanoparticles in all kinds of purposes, and in most cases the ligands
influence the properties of the nanoparticles, more or less. According to the backbone of this issue, let us begin with those of noble metal nanoparticles.
At a fundamental level, optical absorption spectra provide information on the
electronic structure of small metallic particles. The absorption spectra of many
metallic nanoparticles are characterized by a strong broad absorption band that is
58
absent in the bulk spectra. Classically, this giant dipole (or surface plasmon) band is
ascribed to a collective oscillation of the conduction electrons in response to optical
excitation. The optical spectra of gold particles with diameters in the range of 1 to
3.4 nm in aqueous solution are shown in Figure 3.7 [27]. For the entire samples one
can find the continuous background rising toward higher energy. The Mie scattering
from the nanoparticle suspension is considered to be responsible for the particles
smaller than 2 nm in the solutions presenting orange-brown, and no additional
absorption peak appears around 500 nm. This result can also be found in the works
of Fauth et al. [81] and Duff et al. [82].
The spectra are normalized arbitrarily at 800 nm and offset vertically. The solutions are all made in distilled water at a concentration of 86 g/mL.
When the particle size increases to 2 nm [Figure 3.7(d)], superimposed on the
background, a broad surface plasmon band around 500 nm occurs that is attributed
to a collective oscillation of conduction electrons in response to optical excitation
[83]. This band becomes more apparent for the particles with average diameters of
3.4 nm [Figure 3.7(e)]. A similar phenomenon has been reported by Robert L.
Whettens group. Based on an optical study using long-chain thiolate-modified gold
nanoparticles in a similar size range (from 1.4 to 3.2 nm), they reported that the surface plasmon band was essentially unidentifiable for crystallites of less than 2.0 nm
effective diameter [84].
(a)
100 nm
(b)
80 nm
Figure 3.6
Three-dimensional superlattice structure of 7-nm cobalt nanoparticles. (Source: [79].)
59
1.6
Absorbance
1.2
0.8
e
d
c
b
a
0.4
0.0
300
400
500
Wavelength
600
700
800
Figure 3.7 UV-visible spectra of Au particles of different sizes: (a) 10.2, (b) 10.8, (c) 12.8, (d)
19.4, and (e) 33.6 . (From: [27]. 1999 American Chemical Society. Reprinted with permission.)
In Whettens groups investigation, they measured the optical absorption spectra

of a series of nanocrystal gold molecules such as larger, crystalline Au clusters with
effective core diameters ranging from 1.4 to 3.2 nm (constituted of ~70 to ~800 Au
atoms), passivated by a compact monolayer of n-alkylthiol(ate)s, across the electronic range (1.1 to 4.0 eV) in dilute solution at ordinary temperature. After a series
of designed experiments, the conclusions can be summarized as follows [84]: With
decreasing core mass (crystallite size), the spectra uniformly show a systematic evolution. Concretely, (1) the so-called surface plasmon bands are broadened until the
effective diameter of crystallites is less than 2.0 nm, (2) the emergence of a distinct
onset for strong absorption near the energy (~1.7 eV) of the interband gap (5d
6sp), and (3) the appearance in the smallest crystallites of a weak steplike structure
above this onset, which can be interpreted as arising from a series of transitions from
the continuum d band to the discrete level structure of the conduction band just
above the Fermi level. The classical electrodynamic (Mie) theory, based on bulk optical properties, can reproduce this spectral evolutionand thereby yield a consistent
core sizingonly by making a strong assumption about the surface chemical interaction. Quantitative agreement with the spectral line shape requires a size-dependent
offset of the frequency-dependent dielectric function, which may be explained by a
transition in electronic structure just below 2.0 nm (~200 atoms).
After the discovery of methods for preparing gold nanoparticles by Dubois and
Nuzzo in 1992 [32], which has overcome the obstacle of a too broad size distribution limiting the theoretical and successful experimental investigation into the optical properties of the gold clusters, the subsequent discovery and increasingly wide
exploration of the highly stable Au:SR nanocrystal systems over the years has established the stable existence and many properties of these systems, including the
points presented below [84].
They are charge-neutral entities comprising a compact crystalline core of
close-packed metal atoms and a dense mantle of straight-chain groups, which can
60
be prepared quantitatively by at least two distinct methods [19, 32], effective across
the ~1.4- to 3.5-nm core diameter range (~70 to 1,000 atoms). Their surface properties are determined by the surfactant tail group (usually methyl, CH3), and hence
the interactions with external agents (e.g., solvents, strong acids, or bases) and with
each other is weakened; the surfactant groups (RS) initially adsorbed can later be
exchanged with excess thiol (RSH) in solution [8587]. A small metal particle surrounded by a condensed dielectric medium [88] governed the electronic properties.
It is notable that the raw, as-synthesized samples have been shown by mass spectrometry to contain enhanced abundances at certain sizes, which is probably attributable to the filled structural shells of Au atoms. The components of these mixtures
can be fractionated from one another according to size and then handled as purified
molecular substances in various manners [87, 89]. Realistic simulations [90] and
X-ray diffraction patterns obtained on selected samples provided considerable
insight into the structure of these assemblies and their interactions, which is consistent with patterns calculated from theoretically generated structures. Whetten et al.
[91] in 1997 found methods to produce and isolate smaller nanocrystals (1.4 to 1.7
nm) in large quantities, which allows one to obtain a rather complete picture of the
evolution of the optical properties of gold nanoparticles.
Based on the achievement mentioned above, the scientists are able to investigate the properties, especially the optical ones, at will. The spectra shown in Figure
3.8 are arbitrarily normalized at the high-energy end and are presented superimposed, without risk of confusion, based on the fact that the relative spectral intensity
at 2.5 eV decreases monotonically with mass (core size) for each series shown. Furthermore, Figure 3.9 suggests that chain lengths for a given core mass have nearly no
influence on the spectra, while at smaller sizes (~1.7 nm) they can be very sensitive to
sample purity [Figure 3.9(b)].
Cursorily, the evolution of the optical spectrum of Au particles with diameters
ranging from 1.4 to 3.2 nm is so simple that it could even be described as that which
is already known about Au from spectroscopic investigations of unfractionated
samples of nanometer-scale particles in glasses, in solution, and from the smaller
cluster compounds [92, 93]. However, the use of well-fractionated samples of gold
nanocrystals, grown in a strongly etching environment [89], allows one to distinguish certain features that had not been noticed previously:
There is an onset of strong absorption located near 1.7 eV, where it behaves
increasingly distinctly at smaller nanocrystal sizes. These show a clean break
from the preonset behavior occurring at the energy of ~1.6 eV, which can be
attributed to the interband (d sp) absorption edge, as shown in the semilogarithmical scale in Figure 3.10.
A weak, steplike structure occurs, beginning with the onset mentioned earlier,
in the spectra of all the smallest clusters studied. It is clearly evidenced in the
differential (or log-differential) spectral intensities plotted in Figure 3.11 (and
its inset), which transforms the weak steps into a series of distinct peaks and
shoulders. These also bring out the onset structure (1.7 eV) as a first peak.
For these effects to be mastered and understood, one should acquaint oneself with
the elementary facts of the electronic structure of close-packed Au, as are confirmed
61
also to apply to Au clusters by cluster-beam experiments [94]. Now we discuss them

in general. In fact, there are two mechanism that originate the optical absorption
intensity (in the 0- to 8-eV range) in Au. First, common to all metals are the
intraband transitions occurring within the broad conduction band; as far as Au is
concerned, it comes mainly from the 6s1p hybridized atomic orbitals, whose onset
is at zero frequency (or above the Kubo gap in small particles [95]). This leads to a
quite weak absorbance, increasing as the square of the frequency until it approaches
that of the surface plasmon frequency. For conduction electron densities of Au
(~60/nm3), it should appear near 5 eV when the particles have a spherical geometry.
The second interband transitions occur between the 5d10 band and the unoccupied
states of the conduction band, which have an onset at the energy difference between
the highest point in the narrow, flat d band and the lowest unoccupied level of the
conduction band (i.e., at the Fermi level). In bulk Au, this first interband excitation
takes place at the X point of the first Brillouin zone, with energy of 1.7 eV. The substantial d p character of these transitions gives rise to very strong absorption and
sequentially to the colors of Au surfaces, thin films, and nanoparticles. The
researchers consider it to be a truth that the high polarizability of the 5d10 cores
(a)
C6
0
(b)
C12
0
(c)
C18
0
1.5 2.0 2.5 3.0 3.5 4.0

Energy (eV)
Figure 3.8 Optical absorption spectra for dilute solutions of several purified gold molecules passivated by (a) hexyl-, (b) dodecyl-, and (c) octadecylthiolates. The spectra are scaled to unity at 4 eV
and are compared to an aqueous solution of commercial colloidal Au particles (dotted lines) of
9-nm mean size. The peak amplitude (near 2.5 eV) descends with the metallic core diameters: (a)
3.2, 2.5, 2.4, 2.2, 2.0, and 1.7 nm (SC6 passivant); (b) 2.5, 2.4, 2.2, 2.1, 2.0, and 1.7 nm (SC12
passivant); and (c) 1.7 and 1.4 nm (SC18 passivant). (From: [84]. 1997 American Chemical Society. Reprinted with permission.)
62
(equivalent to a dielectric constant near 10) results in a second effect, which is the
large redshift of the collective resonance to its observed location (2.4 eV), with the
unusual results that its intensity is dominated by the interband transitions, its existence by the intraband transitions, and its location by the latter but modified
strongly by the polarizable Au+ ion cores. It seems that the breadth and proximity of
this resonance compared to the interband transition onset will make it more difficult
to distinguish the interband absorption edge. Fortunately, Taylor et al. [94] gained
the photoemission spectra on mass-selected cold AuN- beams, thereby demonstrating that this interband energy, which is located at just below 2 eV, was defined
clearly and exactly for clusters of N = 21 to more than 200 atoms. As a result, its
location is probably constant for all close-packed Au clusters, in spite of the fraction
of atoms at the surface. Based on these facts, the onset feature observed near 1.7 eV
can be ascertained without ambiguity to the first interband transitions, that is, to
transitions from the highest occupied d orbitals to the lowest unoccupied level(s) of
the conduction band.
Because no such structure has previously been ascertained in condensed Au, it
is not easy to propose an interpretation of the discrete steplike structure (the peaks in
Figure 3.11). However, by elimination methods, the evidence points toward the reason that it results from the pattern of unoccupied energy levels located just above the
Fermi level, that is, from the quantum size effect in the conduction band. Each step
would then represent a newly accessible channel in the sparse conduction bandlevel
structure for transitions from the quasicontinuum of d-band levels.
Scientists prefer to address the applicability of various size-dependent corrections to the optical properties derived from bulk Au in a quantitative analysis of the
optical spectra of passivated gold nanocrystals. The effects of surface scattering
(mean free path correction), surfactant effects on the core electron density, and
(a)
(b)
1.5 2.0 2.5 3.0

Energy (eV)
3.5
Figure 3.9 Optical spectra comparing nanocrystals of like metallic cores. (a) Spectra (normalized
at 4 eV) from 2.5-nm-diameter clusters passivated with hexyl (solid line) and dodecyl (dashed). (b)
Spectra (offset for ease of viewing) of 1.7-nm clusters passivated by (top to bottom) SC6, SC12,
and SC18. (From: [84]. 1997 American Chemical Society. Reprinted with permission.)
63
quantum size effects are taken into account. Now it is time to discuss the basic Mie
theory.
The essential assumption of Mies theory of (linear) optical absorption by
small particles is that the particle and its surrounding medium are each homogeneous and describable by their bulk optical dielectric functions. Briefly, by analyzing Maxwells equations for this geometry one can obtain an expression for the
absorption cross section, which is a sum over electric and magnetic multipoles
(spherical vector harmonics and Legendre polynomials). In the case that the size of a
particle is much smaller than the wavelength of the exciting radiation, the absorption is dominated by the dipole term, with a cross section () given by [96]
( ) = 9 m3 / 2 V0
2 ( )
c ( ) + 2 2 + 2 ( )
m
1
2
(3.4)
In this expression, is the frequency and c is the speed of light, m is the dielectric
constant of the embedding medium (assumed to be frequency independent over the
spectral range of interest), V0 is the volume of the absorbing particle, and 1() and
2() are the real and imaginary parts of the frequency-dependent dielectric constant of the absorbing solid.
According to the bulk optical constants [97] extracted by cubic spline fit,
Whetten calculated the bulk dielectric function for Au and the corresponding
absorption spectrum based on (3.4). The spectrum (Figure 3.12), calculated assuming m = 1, shows a giant dipole resonance peaking near 2.3 eV, which is consistent
with the vanishing of 1() + 2m . From (3.4) one can see that the spectral features
10.00
1.00
0.10
0.01
1.0
1.5
2.0
3.0
2.5
Energy (eV)
3.5
4.0
Figure 3.10 Optical spectra (offset for ease of viewing) of diverse selected fractions replotted on a
semilogarithmic scale (top to bottom): 9-nm colloidal gold and 3.2-, 2.5-, 1.7-, and 1.4-nm
nanocrystal gold molecules. The vertical dashed line marks the onset of the interband absorption
just above 1.6 eV. (From: [84]. 1997 American Chemical Society. Reprinted with permission.)
64
dA/d
0
1.5
1.5
2.0
2.5
Energy (eV)
2.0
2.5
3.0
3.5
Figure 3.11 Derivative (with respect to wavelength , dA/d ) of the optical absorption of diverse
selected fractions (top to bottom): 2.6-, 2.4-, 2.2-, 2.0-, 1.7-, and 1.4-nm-diameter nanocrystal
1
gold molecules. The inset shows the logarithmic derivative (A dA/d ) for the smallest of these.
(From: [84]. 1997 American Chemical Society. Reprinted with permission.)
are size independent and the particle dimension acts only as a volumetric scaling factor. To obtain a size dependence for metal particles, one must decompose the dielectric function into two terms: an interband (IB) contribution, accounting for the
response of 5d electrons, and a free-electron contribution (Drude, D) [98] from the
electrodynamics of the nearly free conduction electrons [99]:
1 ( ) = 11B ( ) + 1D ( ),
1 ( ) = 1
2p
2 + 20
2 ( ) = 21B ( ) + 2D ( )
2D ( ) =
2p 0
2 + 20
(3.5)
(3.6)
Here, p is the frequency of the plasma oscillation of free electrons expressed in

terms of the free-electron density N, the electron charge e, and the effective mass m:
2p = Ne 2 / m
(3.7)
which corresponds energetically to 8.89 eV for gold and 9.08 eV for silver, respectively [98]. The term 0, which usually takes a magnitude on the order of hundredths of an electron volt, acts as the deputy of the frequency of inelastic collisions
(electron-phonon coupling, defects, impurities) of free electrons within the metal.
From Figure 3.12 we can see the intuitive separation of susceptibilities of bulk fcc
Au. The variation in the 1() is free electron in nature below 2.4 eV, although offset
by the large, positive, and near-constant interband contribution. A significant inflection appears near 2.4 eV. The 2() function is dominated by the free-electron term
65
10
11B
ID
10
20
(a)
30
40
50
6
21B
5
4
(b)
3
2
2
1
2D
0
1
1.5
2.5
3.5
4.5
Energy (eV)
Figure 3.12 Decomposition of the experimental dielectric functions (solid lines) of bulk fcc Au
into free-electron (Drude) and interband (5d f 6sp) contributions: (a) real and (b) imaginary parts.
The onset of significant interband absorption [dotted curve in (b)] can be seen near 1.8 eV. (From:
[84]. 1997 American Chemical Society. Reprinted with permission.)
below 1.7 eV and by the interband contribution above 2.4 eV. To introduce size
effects, one should assume that as the size of the particle diminishes, the rate of scattering from the particle surface (s) begins to greatly exceed the bulk scattering rate
0 [99]. The surface scattering rate is described in terms of the Fermi velocity (F =
8
1.410 cm/s for gold and silver) and particle radius [98]:
s = A F / R
(3.8)
This expression can be explained as a limitation on the mean free path of the free
electrons by the particle sizes. The coefficient A is of the order of unity, and its
meaning is not sufficiently identified [100]. It takes the value unity if the scattering is
assumed to be isotropic, three-fourths if diffusive, or zero if elastic. When other factors such as electron density at the surface, the effect of the interface, the anisotropy
of particle, and quantum mechanical computations are incorporated into the theory, values from 0.1 to above 2 can be deduced and theoretically justified [6]. To
identify dielectric functions of the particles, one assumes that the interband contribution (Figure 3.12) is the same as that of the bulk, but that free-electron contributions for small particles use s in place of 0 in (3.6).
66
Assuming that the scattering rate in nanomaterials is the same as that in the
bulk, the researchers calculated the size evolution of the optical absorption spectrum
and compared the results with those obtained using this mean-free path correction
(A = 1). The conclusion can be described as follows.
The damping and redshift of the giant dipole band are clearly evident. The
redshift in the position of the plasmon band is the result of the correction to 1,
which is not negligible at smallest sizes [99].
Whetten and coworkers attempt to fit the measured absorption coefficients
recorded on SC12 surfactant with Mie theory and simple MFP correction (3.4) to the
Drude terms in the dielectric function. The fits failed to describe the broadening of
the surface plasmons and their positions. Nor did they observe a relative increase in
low-energy absorption, accompanying the quenching of the surface plasmon, which
is demanded by the functional behavior in the mean-free path model (Figure 3.13).
Enlightened by Hengleins work [101], Whetten and coworkers take that
adsorbates can significantly affect the metals electronic properties, both for bulk
surfaces and in small particles as the second correction. The dependence in (3.7) of
the plasmon frequency (p) on the free-electron density N allows for a simple
phenomenological description of the adsorbate effect. It is possible that they provide
or withdraw additional electron density at the interface based on the facts that
thiol(ate)s are intimately bound to the particle surface. In the case of flat gold surfaces with self-assembled thiol monolayers on them, approximately 0.2 nm2 of surface supports one thiol [102], and the coverage density is increased in small particles
because of curvature introduced by edges and vertices [90]. When correction is
0
1.0
1.5
2.0
2.5
3.0
3.5
4.0
Energy (eV)
Figure 3.13 Absorption spectra predicted by Mie theory for particles of decreasing diameter (top
to bottom): 40 (bulk), 3, 2, and 1 nm. All curves were normalized to unity at 4.1 eV. The size
dependence results entirely from mean-free path corrections to both real and imaginary dielectric
functions. (From: [84]. 1997 American Chemical Society. Reprinted with permission.)
67
employed, taking as adjustable parameters the number of electrons donated (or

withdrawn), the core size, and the thiol surface coverage density, one can obtain the
better fit only by adding the assumption that the surfactant groups donate (or withdraw) electrons. In fact, based on the assumption that one electron is donated per
RS chain adsorbed, researchers indeed got reasonable values for the extracted core
dimensions and surfactant packing densities.
Unfortunately, when these modifications are utilized to explain the abnormally wide or depressed collective oscillation band, which resists being fitted with
simple corrections even after our introducing hypothetical cluster-size distributions,
incorporating a Lorentz term for outer core electrons, or invoking Kubos quantum
mechanical correction to the relaxation time [43], the loss is the only thing that is
obtained. Huffman and Henglein together with their coworkers suggested that the
passivating layer provided a dielectric coating on the surface of the sphere, amounting to a change in the dielectric constant of the medium [3, 103]. Another proposed
modificationsupposing that the outer surface gold atoms bound to the thiol(ate)
lose their metallic character and are effectively removed from the metallic core,
leaving its electron density unchangeddoes no better. In fact, both of these modifications make no improvement to the fit of the patterns from calculations and
experiments.
However, regardless of the flaw of the specific theoretical foundation as discussed above, it is possible to reproduce the spectra mathematically by modifying
the dielectric functions, although this must be done in a way that satisfies certain
constraints (sum rule and Kramers-Kronig relations). For example, a much better fit
is obtained simply by adding an energy-independent (offset) correction to the optical constants, and this suggests a transition in the electronic structure of Au at a size
just below 2.0 nm. In 1996, Kreibig et al. [104] suggested that an abrupt change in
the experimentally determined dielectric constants for small gold particles took
place at approximately 3-nm diameter. Whetten et al reported that their results
placed this change in the electronic structure of thiol-passivated gold particles precisely where a transition in the atom-packing structure had recently been found
[27], suggesting that these may be interrelated [84].
3.3.2
Magnetic Properties of Metallic Nanoparticles
When the magnetic properties are talked about in terms of magnetic metallic
nanoparticles, the most noteworthy point of the origin is that the volume is small
and the surface/bulk ratio is big. Because of the small volume, the thermal fluctuation energy of the magnetic moment can compete with the magnetic barrier energy
and even overcome it. Hence, the magnetic moment can rotate freely despite the
magnetic barrier energy. In the most cases, the saturation magnetization is smaller
than the bulk counterpart, and it is attributed to the surface/volume ratio and the
situation of the surface.
In general, there are two basically points in the nanoparticles that are different
from those of their bulk counterpart. First, and also most important, when the particle size is smaller than a certain critical value, they present very different behavior
from their bulk counterpart. Concretely, at room temperature, they exhibit zero
coercive field and no remanence, but they do saturate and the slope of the M-H
68
curve varies continually before it reaches the saturation state. Second, the measured
average moment per molecule (atom, in the case of a metallic nanoparticle) usually
is smaller than the one in bulk materials. The loss in magnetization corresponds to a
partial quenching of the magnetic contribution of the surface. The fact that the magnetization of the surface layer is not completely quenched is explained by a partial
coverage of the surface, in good agreement with the bulkiness of the ligands. Before
the discussion of the magnetic properties, we introduce some terms that are needed
when we talk about the magnetic properties of nanoparticles.
Superparamagnetism. At room temperature, the nanoparticles usually exhibit

zero coercive fields and no remanence, but they do saturate and the slope of
the M-H curve varies continuously before it reaches the saturation state. So
they are considered to be super paramagnetic and the magnetic properties are
called superparamagnetism.
Blocking temperature. As mentioned above, the nanoparticles are superparamagnetic at room temperature. However, when the measurement is taken at
low enough temperature, either the coercive field or the remanence is not zero
any more. The behavior can be fully attributed to ferromagnetism. So there
exists some temperature at which the ferromagnetism begins to take the place
of superparamagnetism. That is the blocking temperature.
Following are the achievements obtained to date. As always, the statements are constructed based on the outstanding work from different groups.
In the work reported by Gaos group [79], magnetic properties of the ~7-nm
cobalt nanoparticles deposited on highly oriented pyrolytic graphite (HOPG) substrate are measured by SQUID using a standard airless procedure. The magnetization as a function of the temperature in a 10-Oe field between 5K and 300K
determines the blocking temperature using a zero-field cooling (ZFC) procedure.
Figure 3.14(a) shows the typical result for magnetic NPs. One can see that below the
critical size at which a particle becomes a single-domain magnet and is small
enough, the nanoparticles display superparamagnetism [106] at high temperature.
From the concrete measurement data, the blocking temperature (Tb) is 92K. The
broad transition from superparamagnetism to ferromagnetism shown in Figure
3.14(a) around 92K is probably due to the magnetostatic particle interactions in the
close-packed arrays. The blocking temperature should roughly satisfy the
relationship
Tb = KV / 30kB
(3.9)
where K is the anisotropy constant, kB is Boltzmanns constant, and V is the average

volume of the particles. With the knowledge of the blocking temperature and the
4
3
particle size, the anisotropy constant is 2.1 10 erg/cm for the Co particles, which
6
3
is smaller than that of the bulk fcc cobalt (2.7 10 erg/cm ).
Figures 3.14(b) and 3.14(c) show the hysteresis loop of the cobalt NPs powder
recorded at 250K and 10K, respectively. Cobalt NPs show no remanent magnetization in their magnetization field data at 250K. As discussed above, the superparamagnetism behavior was observed at room temperature. While at 10K, the remnant
69
M (emu, normalized)
M (emu/g)
20
100
150
Temperature (K)
(a)
50
(b)
200
250
(c)
10
0
10
250K
20
60
(d)
10K
(e)
M (emu/g)
40
20
0
20
40
10K
10K
60
20
10
H(kOe)
10
10
10
20
H(kOe)
Figure 3.14 (a) ZFC magnetization versus temperature of cobalt NPs showing a blocking temperature (Tb) of 92K. Hysteresis loop of powder of cobalt NPs compacted into a capsule obtained at (b)
250K and (c) 10K. Hysteresis loop obtained at 10K: (d) diluting cobalt NPs with wax and (e) cobalt
NPs deposited on HOPG and dried under N2 to prevent oxidation. (Source: [79].)
magnetization Mr is about 1.5 emu/g, the coercive field Hc is 163 Oe, and the magnetization at saturation (Ms) is estimated to be only 14.0 emu/g (the estimation is
based on an extrapolation of curves of H/M versus H).
The noteworthy point of this work is the measurement conducted in different
states of the cobalt nanoparticles. Yang et al. diluted the Co NPs in wax with a mass
ratio of cobalt nanoparticles:wax equal to 1:4. Figure 3.14(d) shows the hysteresis
loop of diluted particle powder, and a clear change in the shape of the hysteresis
loop can be found. The Mr reaches 7.3 emu/g, Ms reaches 59.6 emu/g, and Hc
increases from 163 to 600 Oe in comparison with the cobalt nanoparticles in powder states. The hysteresis loop of ordered arrays of cobalt nanoparticles on HOPG
substrate is shown in Figure 3.14(e). The Mr reaches 12.6 emu/g and Hc increases to
790 Oe. However, the improvement of Ms is not obvious compared to that of the
particles diluted with wax, which is 61.6 emu/g. These values are lower than those
70
obtained from the bulk phase. Because TEM images taken over large areas of the
sample show no evidence of the coalescence, the observed changes cannot be attributed to the coalescence of the cobalt NPs. They suggested that the exchange coupling between adjacent particles should account for explanations for the change in
magnetic properties. The dipole coupling enhancements are attributed to the longrange order of the two-dimensional lattice and collective flips of the magnetic
dipoles.
In the work of Catherine Amiens [6467], she and coworkers discussed the magnetic properties of the cobalt NPs they synthesized. The most striking result is the
high differential susceptibility observed in high fields and leading to a very low magnetization at 5T in a certain sample (namely, sample 1). The fact that the NPs adopt
structures very different from the bulk ones, even if it may lead to strong changes in
magnetic anisotropy, should not account for this strong decrease in magnetization.
Indeed, body-centered-cubic (bcc), fcc, and hcp phases present very close values of
magnetic moment per atom (1.6 to 1.7 B/Co). It is also the same in -Co, where the
value determined is close to 1.7 B/Co, or in polytetrahedral arrangements, where no
magnetization reduction could be evidenced, whereas the atom packing is very different from the bulk one [52, 53]. Either partial oxidation of the NP surface or a
strong damping of the magnetic moment of the surface atoms introduced by coordination of the ligands or other chemical species at their surface could then result in
this magnetization damping. Either through a strong alteration of surface anisotropy or via the formation of a diamagnetic surface layer, a strong coordination
effect would also be responsible for the fact that the magnetization is difficult to saturate. Oxidation can be ruled out by the given XANES and EXAFS spectra. This
sample was synthesized under a di-hydrogen pressure, whereas another sample
(namely, sample 2) was not. In fact, surface hydrides are expected to be present at
the surface of the particles, as recently demonstrated for Ru NPs [104]. One can thus
question the effect of hydrogen adsorption on the magnetic properties. However, in
Selwoods book one can find experiments showing that hydrogen adsorption
induces only a small decrease of magnetization in the case of cobalt [107]. Furthermore, as Amiens et al. demonstrated in 1998, NPs prepared in the same conditions
of hydrogen pressure but stabilized by poly-(vinylpyrrolidone), displayed magnetic
properties similar to those of free-standing Co NPs [52]; hence, the possible role of
coordinated hydrides in the low Ms value found for sample 1 was ruled out. However, the ligand used during the synthesis of sample 1 can strongly interact with
hydrides coordinated to transition metals [24]. In this particular case, the formation
of hydrido aluminate species at the NP surface should take place, which could then
transform into tricoordinated surface alkyl aluminum species. Formation of surface
alkyl radicals may also be envisaged, in agreement with the short distance observed
by EXAFS. Hence, the surface of the NPs should then be regarded as electronically
depleted. As is well known, electron-withdrawing ligands such as carbon monoxide
deprive the surface atoms of their magnetic properties [108]. In the assumption of
Amiens et al., a similar phenomenon takes place in sample 1 and accounts for the
observed low magnetization.
The loss in magnetization corresponds to a partial quenching of the magnetic
contribution of the surface. The fact that the magnetization of the surface layer is
3.4 Application of Metal Nanoparticles in Biomedicine
71
not completely quenched is best explained by a partial coverage of the surface, in

good agreement with the bulkiness of the ligands.
Now, we have a grasp on the primary properties of the metal nanoparticles and
can move into the bioapplications of them.
3.4
Application of Metal Nanoparticles in Biomedicine

3.4.1
3.4.1.1
Biomedical Detection Using Novel Metal Nanoparticles

Au Nanoparticles
Functional nanomaterials that are designed to perform a reaction are of intense

interest because of their potential uses in medical diagnostics [109], drug delivery
[110], and catalysis [111, 112]. To be useful and biocompatible for biomedical
application, the nanoparticles must be coated by and/or linked with suitable molecules, and the most well-understood method is the utilization of place-exchange
reactions, which are based on displacement of surface coatings. Regarding the realization of the bioapplication of the NPs, the technique that allows us to modify the
surface of the NPs must play the determinative role. So we introduce this even more
than the idiographic bioapplication. The employment and generality of these reactions for the modification of Au nanoparticles was first described by Ingram et al.
[113], who produced chemically useful particles by replacement of the monolayer
ligands with -functionalized alkanethiols. A significant drawback to this method
is the need to synthesize individual thiolated ligands for the aim at insertion into the
monolayer. The inclusion and tailoring of functional moieties has been an aim in
small-molecule organic chemistry for decades, and as a result, a huge number of
reactions have been developed to convert, for instance, alcohols into carboxylic
acids and halides into alkenes [114] or to combine them with molecular species
using amide or ester condensation chemistry [115]. These reactions have all been
employed for the modification of Au nanoparticles surface structure and chemistry
[116]. However, these chemistries are not compatible with every desired application, and the result is that the development of additional routes in general toward
nanoparticle functionalization is still necessary for their use in emerging
nanotechnologies. Molecular species can be conjoined by 1,3-dipolar cycloaddition
reactions [117], which were first described by Huisgen [118] and have recently
gained a revisit with the advent of click chemistry.
In the latter method, azide-containing based on the discussion from triazole
coupling reactions could analogously be utilized as a general method for the
functionalization of metal nanoparticle surfaces. Elizabeth Williams et al. [119]
designed a system as a general route for functionalization of Au nanoparticles.
Based on the facts that only mild reaction conditions are required and that the
extreme selectivity toward molecules bearing azides and alkynes prevents unwanted
side products, this reaction scheme is a particularly effective design. Figure 3.15
shows the general synthetic strategy for triazole functionalization of nanoparticles.
Let us tell about the synthesis of a series of redox-active, fluorescent, and
solubilizing species and the use of click chemistry as a facile route toward
functionalization of monolayer-protected Au nanoparticles.
72
(a)
Br
(b)
N
=N= N
=N= N
(c) =
R
N
=N= N
=
R
O
Figure 3.15 (a) Br(CH2)11SH in DCM, 60 hours at room temperature; (b) 0.25 M NaN3 in
DCM/DMSO solution, 48 hours; and (c) R = propyn-1-one derived compounds as in scheme 2, 24
to 96 hours in dioxane or 1:1 hexane/dioxane. (From: [119]. 2006 American Chemical Society.
In the case of synthesis of 1-Pyren-1-yl-propyn-1-one (Pyr), an amount of

pyrene-1-carboxaldehyde was set in a round-bottomed flask together with anhydrous tetrahydrofuran (THF) with ethynylmagnesium bromide as solute. From the
beginning to the end of the process, the reaction system was protected under nitrogen. The reaction was quenched with saturated aqueous ammonium chloride after
the solution was allowed to be stirred overnight. Finally, the product was extracted
with diethyl ether. The combined organic layers were dried over sodium sulfate, and
the ether was removed. After the product was dissolved in 50 mL of anhydrous acetone, Jones reagent was added dropwise under stirring until a red color occurred.
After the reaction was quenched with isopropyl alcohol, the solution color turned
green. An excess of saturated aqueous sodium metabisulfite was added in, then the
acetone was removed by rotary evaporation. After the extraction and purification,
an intense yellow solid was obtained. The synthesis of 1-Anthracen-9-ylpropyn-1-one (An), Propynoic Acid 2-[2-(2-Methoxy-ethoxy) ethoxy]ethyl Ester
(PEG) and Propynoic Acid Phenylamide (Ani) is similar except for a little change or
modification and the use of some new reactant instead [119].
They prepared the modified Au particles by employing the two-phase synthesis
method of Brust et al. [120] with slight modifications. Briefly, HAuCl4 in distilled
water was transferred into toluene using tetraoctylammonium bromide and the
organic phase was isolated. After decanethiol was added, the solution was cooled to
0C and stirred for 10 minutes, after which an aqueous solution containing 380 mg
of NaBH4 was added in. The mixture was stirred for an additional 3 hours before the
organic layer was separated and evaporated; a black waxy solid was produced and
was washed with copious amounts of ethanol.
Imitating the approach employed by Murray and coworkers, Williams et al. dissolved the as-synthesized Au nanoparticles and an amount of BrC11H22SH in DCM
and kept the mixture under stirring at room temperature for 60 hours to perform
ligand exchange with BrC11H22SH. Following standard characterization methods
[121], the NMR and Fourier transform infrared (FTIR) spectra reveal that protective monolayers contain both CH3- and Br-terminated ligands.
To the synthesis of azide-functionalized Au nanoparticles, they dissolved the
Au--Brfunctionalized particles in DCM and then added them into an equal volume of NaN3 in DMSO. After the solution was allowed to remain under stirring for
73
48 hours, they added water and the black organic layer was isolated. They performed reaction of azide-functionalized Au nanoparticles with alkynyl derivatives
using a procedure that can be described as follows: N3-functionalized particles and
the alkynyl-modified compound (Figure 3.16) were codissolved in dioxane or hexane/dioxane and stirred for 24 to 96 hours. Under vacuum, the solvent was removed
and ethanol was utilized to remove any unreacted alkynyl derivative. Finally, the
particles were dried and then redissolved in DCM. Additionally, Au nanoparticle
samples could be decomposed using standard disulfide-forming reactions. For testing the functionalization approach, reactions involving triazole ring formation were
conducted using small Au particles.
Because its length sufficiently stabilizes the particles from aggregation but is not
so sterically hindered as to prevent ligand exchange, decanethiol served as the surface ligand in the synthesis of monolayer-protected Au clusters.
One can see in the representative TEM image in Figure 3.17 that the Au nanoparticles are spherical and have an average diameter of about 1.8 nm. For the purpose of replacing a fraction of these ligands with Br-terminated undecanethiol
ligands, the decanethiol-stabilized Au particles were then stirred in a solution containing BrC11H22-SH. By means of NMR and FTIR spectra, the replacement was
confirmed. In the reaction of the Br termini, in the way that nucleophilic substitution with NaN3 was used to append azide functionalities to the Au nanoparticles, as
shown in Figure 3.17(b), the size and shape of the resultant particles were not
affected. According to the FTIR and NMR spectra, the ligand replacement with
BrC11H22-SH and subsequent reaction with NaN3 and the resultant Au
nanoparticles containing mixed monolayers that are 44% CH3- and 52% N3-terminated alkanethiol ligands were confirmed.
In Williamss work, they also conducted the triazole functionalization of Au
nanoparticles, and further particle functionalization through 1,3-dipolar cycloaddition reactions (i.e., click chemistry), by fusing ethynyl- and azide-bearing
O
O
Fe
NO2
Fc
NB
Pyr
O
O
CH3
3
HN
O
An
PEG
Ani
Figure 3.16 Scheme 2. Propyn-1-one compounds for attachment via triazole ring formation.
(From: [119]. 2006 American Chemical Society. Reprinted with permission.)
74
# of Particles
160
120
80
40
0
0
# of Particles
50
40
30
20
10
0
0 1 2 3 4 5 6
Size (nm)
(a)
2
4
Size (nm)
(b)
Figure 3.17 (a) Representative TEM images of synthesized C10H21SH-modified Au particles and (b)
azide-functionalized Au nanoparticles. Insets contain particle size distributions. The scale bar is 50
nm. (From: [119]. 2006 American Chemical Society. Reprinted with permission.)
molecules, can be allowed by decorating the nanoparticle surface with N3

functionalities. They utilized six different alkynyl compounds including the alkyne
derivatives of ferrocene (Fc), nitrobenzene (NB), pyrene (Pyr), anthracene (An),
poly(ethylene glycol) (PEG), and aniline (Ani) to demonstrate the utility and generality of this method [119]. After the reactants were connected to the Au nanoparticles, a range of physical, electronic, and spectroscopic properties are
introduced or improved. Aiming at the enhancement of the rate of triazole formation, each was synthesized to include a carbonyl group adjacent to the terminal
alkyne to provide a more electron-withdrawing environment [117, 123].
According to the NMR and FTIR results recorded for the as-synthesized functionalized Au nanoparticles (for the purpose of this chapter, we prefer to ignore the
concrete procedure for preparing samples and the patterns measured).
Other than the ring formation, the surface attachment does not affect the structures of the compounds drastically. In addition, the alkyl stretching vibrations of the
alkanethiolate remain unchanged by cycloaddition, indicating that the protecting
surface monolayer is largely unaffected by triazole ring formation. The complete
reaction that the triple bonds with the terminal N3 groups on the Au nanoparticles to
form a triazole ring rules out binding by intercalation of the ligand into the particle
surface monolayer or the possibility of unreacted alkynyl compounds present as
impurities.
In this approach they also utilized cycloaddition reactions to prepare multifunctional Au nanoparticles by simultaneously stirring the N3-terminated nanoparticles
with several acetylenic small molecules to obtain N3-containing Au nanoparticles,
which was reacted with a solution containing both the Fc and the NB propyn-1-one
species. Analysis based on FTIR results indicates that the multifunctional Au
nanoparticles containing both Fc and NB species linked through the fact that a
triazole ring formed.
75
Quantitative assessment of extent of this functionalization approach has been

carried out following a method that has been previously described [119]. An estimation of the percentages of surface-bound ligands per Au nanoparticle was performed by assuming the following:
1. No ligands are destroyed though side reactions.
2. All ligands are completely cleaved from the surface by means of reaction
with I2. Analysis of peak integrations in the NMR result for the
N3-terminated particles suggests that there are about 1.2 N3-terminated
thiols for every CH3-terminated thiol and an average of ~4% ligands per
particle that are not converted to N3 upon reaction with NaN3 [119].
However, this may result from the fact that a difference in the degree of solubility of the reactants exists and there is no apparent tendency with respect to size or
reactivity for Pyr- or PEG-coupled Au nanoparticles. As far as the role of solvent on
the extent of functionalization is concerned, by repeating these reactions in a range
of polar and nonpolar solvents, it was found that the latter ones are more effective
and have a larger yield.
Since solubility played a primary role when determining reactivity, the researchers have observed that solutions containing both hexane and dioxane would
solubilize both the Au nanoparticles and click compounds. In the click compound
studied, an average threefold increase in the efficiency of the azide conversion to
triazole is the result from addition of the more nonpolar (such as hexane) solvent.
These facts are in good agreement with prior reports, where hydrophobic solvents
better stabilized the nonpolar transition state [124, 125].
The better solubilization of the hydrophobic monolayer on the Au nanoparticle
surfaces by addition of the nonpolar hexane to solution should also possibly
account for this effect.
Sharpless et al. have demonstrated that Cu catalysts can greatly enhance the
rate of triazole formation, particularly in aqueous solutions, which is a prerequisite
[126, 127], so the researchers have also sought a proper catalyst system to increase
the conversion. Because of the hydrophobic habit of the monolayer-protected particles, which made them insoluble in aqueous solutions and prevented the use of the
most commonly employed CuSO4-ascorbic acid system, the use of several organic
soluble catalysts, including bromotris(triphenylphosphinato) copper(I) [128], CuI,
[129], and CuBr/2,6-lutidine [130], seems reasonable. However, in all cases immediate and extensive particle aggregation or decomposition was observed. This phenomenon may be caused by the attractive force between the particle surface and the
organic coating, which may have different charges.
By the use of triazole ring formation as a general method, chemical, spectroscopic, or electrochemical functionality may be appended to metal nanoparticles.
This procedure can be named clicked functionality on Au nanoparticles. Hence,
we take an overall look at the modification of the surface. And the idiographic
application of Au nanoparticles will be introduced, together with Ag NPs, in the
next section.
76
3.4.1.2
Ag Nanoparticles
First, we introduce the useful technology that utilizes localized surface plasmon resonance (LSPR) nanosensors in detecting biological molecules. This novel, nanoscale
development is a refractive indexbased sensing device that relies on the extraordinary optical properties of noble (Ag, Au, Cu) metal nanoparticles [131135].
Briefly, it is based on the nanoscale limit of surface plasmon resonance sensors. In
other words, LSPR, which refers to the ability of the conduction electrons in the
nanoparticle to oscillate collectively [136], induces electromagnetic fields surrounding the nanoparticle, which determine the sensing volume in which refractive
indexbased sensing can occur [135, 137]. Nanoparticles exhibit selective photon
absorption, which can easily be monitored using ultraviolet-visible, since it is well
known that the conduction electrons oscillate collectively to only specific wavelengths of light (UV-visible) spectroscopy. It is well established that the maximum
extinction wavelength max of the LSPR is dependent upon the composition, size,
shape, and interparticle spacing of the nanoparticles. And the dielectric properties of
their local environment (i.e., substrate, solvent, and surface-confined molecules)
[135, 136] also play an important role.
The first nonmodel application of the LSPR nanosensor was reported by Haes et
al. in 2004 [135]. In their work, the LSPR nanosensor underlain by the optical properties of Ag nanotriangles was shown to aid in the understanding of the interaction
between amyloid derived diffusible ligands (ADDL) and the anti-ADDL antibody,
molecules possibly involved in the development of Alzheimers disease. By varying
the concentration of anti-ADDL antibody, a surface-confined binding constant of
3.0 107 M1 for the interaction of ADDLs and anti-ADDLs was measured. Influences of Cr, the nanoparticle adhesion layer, will be shown to be the limiting factor
in the sensitivity of this assay.
In fact, in 2002, they presented the work on the employment of Ag nanoparticles
together with Au nanoparticles in the LSPR procedure. Briefly, it is based on the fact
that triangular silver nanoparticles (~100 nm wide and 50 nm high) have remarkable optical properties and the peak extinction wavelength max of their LSPR spectrum is unexpectedly sensitive to nanoparticle size, shape, and local (~10 to 30 nm)
external dielectric environment. This sensitivity of the LSPR max to the
nanoenvironment has provided the opportunity for developing a new class of
nanoscale affinity biosensors. Utilizing the well-studied biotin-streptavidin system,
the essential characteristics and operational principles of these LSPR nanobiosensors were demonstrated. In this work, a 27.0-nm redshift in the LSPR max
occurred, after the exposure of biotin-functionalized Ag nanotriangles to 100-nM
streptavidin (SA). They measured the LSPR max shift, R/Rmax, versus the [SA]
15
6
response curve over the concentration range 10 < [S] < 10 M. By comparing
the data with the theoretical normalized response expected for 1:1 binding of a
ligand to a multivalent receptor with different sites but invariant affinities, one got
approximate values for the saturation response. The values are Rmax = 26.5 nm
together with the surface-confined thermodynamic binding constant Ka,surf = 1011
1
M for the actual limit of detection (LOD) for the LSPR nanobiosensor, which is in
the low-picomolar to high-femtomolar region. Hence, a strategy for amplifying the
response of the LSPR nanobiosensor with employment of biotinylated Au colloids
and thereby further improving the LOD was carried out. Several control
77
experiments were conducted to define the LSPR nanobiosensors response to nonspecific binding as well as to demonstrate its response to the specific binding of
another protein. They then drew the conclusion that the factors determine the LSPR
procedure could be summarized as follows:
1. Electrostatic binding of SA to a nonbiotinylated surface;
2. Nonspecific interactions of prebiotinylated SA to a biotinylated surface;
3. Nonspecific interactions of bovine serum albumin to a biotinylated surface;
4. Specific binding of antibiotin to a biotinylated surface.
The LSPR nanobiosensor provides a pathway to ultrasensitive biodetection experiments with extremely simple, small, light, robust, and low-cost instrumentation
that will greatly facilitate field-portable environmental or point-of-service medical
diagnostic applications.
In 2003, Riboh et al. published their sequential results in the Journal of Physical Chemistry B [134]. In the later work, the width of the silver nanoparticles was
narrowed to 90 nm. The target was the probe of the interaction between a surfaceconfined antigen, biotin (B), a solution-phase antibody, and antibiotin (AB). An
about 38-nm redshift in the LSPR max took place after exposure of biotin7
6
functionalized Ag nanotriangles to 7 10 M < [AB] < 7 10 M. The experimental normalized response of the LSPR max shift, (R/Rmax), versus [AB] was mea10
6
sured over the concentration range 7 10 M < [AB] < 7 10 M. The similar
comparison as mentioned above led to values for the saturation response, Rmax =
7
38.0 nm, the surface-confined thermodynamic binding constant, Ka,surf = 4.5 10
1
10
M , and the limit of detection (LOD) < 7 10 M. The experimental saturation
response was explained in terms of a closest packed structural model for the surface
B-AB complex in which the long axis of AB, lAB = 15 nm, is oriented horizontally
and the short axis, hAB = 4 nm, is oriented vertically to the nanoparticle surface.
Based on this model, a quantitative response for the saturation response, Rmax =
40.6 nm, is obtained, which is in good agreement with experiment, Rmax = 38.0
nm. By means of atomic force microscopy (AFM), this interpretation was confirmed. Furthermore, they also described the major improvements in the LSPR
nanobiosensor. A series of LSPR nanobiosensor substrates was utilized such as
glass, mica, and a surfactant, Triton X-100, in the nanosphere lithography fabrication procedure. These changes increased the adhesion of the Ag nanotriangles by a
factor of 9 as determined by AFM normal force studies. The improved adhesion of
Ag nanotriangles allows the study of the B-AB immunoassay in a physiologically
relevant fluid environment as well as in real time. These results represent important
new steps in the development of the LSPR nanosensor for applications in medical
diagnostics, biomedical research, and environmental science.
In addition, the Ag nanoparticles can form the nanoscale bioconjugated superstructures together with semiconductor one-dimensional nanostructures, and the
system shows a twofold enhancement of luminescence intensity. Theoretical calculations of the electric field in the cylindrically organized NPs and experimental data
indicate that the enhancement in emission is based on the increase in absorption of
the Ag NP shells in the regime of the collective plasmon resonance. This situation is
78
fundamentally different from the PL (photo-induced luminescence) enhancement in

the Au-NP/NW system. The fundamental importance of these findings is as follows:
1. The results demonstrate that the spatial organization of metal particles has
direct influence on the optical properties as a result of the collective nature of
interactions.
2. The described calculation method can be used to predict properties of
nanoscale superstructures.
3.4.2
Drug Delivery and Biosensing with Magnetic Nanoparticles
Based on the novel properties of nanoparticles discussed above, lots of approaches

emerge that aim at their employment in bioapplications. However, if safety is taken
into account, there still are some obstacles to overcome. Biosuitable magnetite
nanoparticles have been used to date in both experiments in the laboratory and in
clinical applications and industry, so it may be reasonable to introduce the
bioapplication of metallic NPs through that of Fe3O4 nanoparticles. We will introduce a simple, rapid, and ultrasensitive procedure for detection of bacteria in human
blood or blood products using biofunctional magnetic nanoparticles in combination
with a fluorescent probe (a conjugate of vancomycin and fluorescein). Pathogen
contamination in blood products constitutes a great risk to patients (e.g., causing
sepsis), resulted from the fact that the recipients of blood transfusions are usually
critically ill or seriously immunocompromised. Several measures, already advanced,
have reduced the pathogen contamination in blood: The improvement of blood collection and processing (e.g., using a closed system of sterile interconnected plastic
bags to replace an open system of glass bottles) has reduced sepsis significantly
[137], more stringent donor selection and progress in viral-testing technologies have
substantially minimized the incidence of transfusion-transmitted viral infection
[138], and more rigorous skin-disinfection protocol has reduced the rate of bacterial
contamination of blood [139]. Regardless of these improved methods for reducing
pathogen contamination of blood products, the risk of transfusion-associated bacterial infection remains a major problem [140]. This mainly results from bacterial
contamination of platelet concentrates. Because the source of contamination is usually from the skin of the donor [141] or bacteria from the blood of donors who are
asymptomatic carriers [142], it is impossible to completely eliminate bacterial contamination in blood products. In addition, the rate of bacterial contamination
increases with the duration of storage of the platelet [143] because it has been stored
at room temperature. Therefore, it is necessary to monitor the presence of bacteria
in blood components, particularly in the platelets, before transfusion.
In order to generate multivalent interactions [144, 145] and capture the bacteria
from the blood samples, Bing Xu et al. used magnetic nanoparticles [146] as the
platform. Then the enriched bacteria were stained with fluorescent Van. This design
gave an opportunity for the detection of bacteria from the samples within 2 hours
and had sensitivity as low as 10 cfmL1, (the concentration of bacteria in the sample
was confirmed by back titration [147]). This simple strategy has several advantages
over other methods, as follows:
79
1. Magnetic nanoparticles (less than 10 nm in diameter) allow high

performance because of their large surface/volume ratios and easy
attachment to bacteria.
2. Multivalent interactions at the dimension of 10 nm, resembling
immunoglobulin M (IgM), provide high affinity.
3. The combination of magnetic manipulation and fluorescence gets rid of the
need for expensive or complicated instruments and allows implementation
on a portable or handheld device. In addition, the process demonstrated in
this work can be used for detecting other biomolecules or organisms.
Biocompatible magnetic nanocrystals with reactive moieties on the surface were
generated by means of a one-pot reaction and used directly in cancer detection by
being coupled with a specific cancer-targeting antibody. Many reports have demonstrated that the magnetic nanocrystals have potential application in MRI.
In fact, magnetic nanocrystals have shown great potential in biomedical applications (see Figure 3.18) [148152]. In comparison with in vitro applications, in
vivo applications of magnetic nanocrystals are more complicated and challenging
[153, 154]. Researchers have demonstrated that both particle size and surface
chemical structures are critical parameters that determine the blood half-life,
biokinetics, and biodistributions and the opsonization of magnetic nanocrystals
[155, 156]. As far as this context is concerned, there are several bottlenecks in the
synthesis of high-quality magnetic nanocrystals compatible with in vivo applications, which are as follows:
Antibody
Tumor
Before
10 min
6h
10 h
24 h
Figure 3.18 Sketch map of the magnetic nanoparticles serving as biodetection agents. (Courtesy
of Professor Mingyuan Gao, Institut of Chemistry, Chinese Academy of Science.)
80
1. A different surface chemistry for improving the biocompatibility of magnetic

nanocrystals needs to be advanced [157, 158].
2. Reactive moieties on the surface of biocompatible nanocrystals are usually
required for further linking of different bioligands for sophisticated in vivo
applications [153, 154]. However, there are simple and convenient
approaches well documented so far for producing such magnetite
nanocrystals [157, 158].
3. New synthetic chemistry to produce high-quality magnetic nanocrystals is
needed when taking the appropriate surface chemistry for biocompatible
modification into consideration. However, there are only limited examples
of nanocrystal synthetic chemistry being successfully combined with
biocompatible surface modification [159].
By means of coupling the organic-coated magnetic nanocrystals with a cancertargeting antibody (rch 24 mAb), examination of their potential as MRI contrast
agents was carried out. Based on an ethyl-3-(dimethylaminopropyl) carbodiimide
(EDC)mediated reaction, the amidation coupling between rch 24 mAb and
organic-coated magnetic nanocrystals was carried out by Gao et al. Native
polyacrylamide gel electrophoresis was adopted to evaluate the conjugation
reaction.
The T2-weighted magnetic resonance images shown in Figure 3.19 undoubtedly
indicate that cells treated with rch 24 mAb conjugates [Figure 3.19(a)] exhibit a
much faster signal decay, that is, a much shorter proton T2 relaxation time, which
suggests that the magnetic nanocrystals can effectively target and be bound to the
cancer cells via rch 24 mAb. In contrast, no obvious difference can be seen between
the control experiments [Figure 3.19(b),(c)] and the untreated cancer cells [Figure
3.19(d)], which suggest that the rch 24 mAb conjugates have an excellent binding
selectivity to the cancer cells.
d
25ms
75ms 125ms 175ms
Figure 3.19 Left: Photograph of 5% polyacrylamide separating gel stained by coomassie blue.
The four lanes from left to right are filled with Fe3O4-(rch 24 mAb) conjugates, a mixture of Fe3O4
and rch 24 mAb, Fe3O4 nanocrystals, and rch 24 mAb, respectively. Right: T2-weighted MR images
of CEA-positive LS 180 cells treated with (a) Fe3O4-(rch 24 mAb) conjugates, (b) Fe3O4-irrelevant
antibody conjugates, and (c) Fe3O4@PEG-COOH nanocrystals. (d) Untreated LS 180 cells shown for
comparison. The echo time used to acquire the images was (left to right) 25, 75, 125, and 175 ms.
(Courtesy of Professor Mingyuan Gao, Institute of Chemistry, Chinese Academy of Science.)
81
Based on the success of in vitro experiments, the rch 24 mAb conjugates were
used in the subsequent in vivo experiments aiming at detecting human colon carcinoma xenograft tumors implanted in nude mice at their proximal thigh region.
The conjugates were injected through a tail vein into a group of mice carrying
tumors, and the magnetic nanocrystals were intravenously injected into another
group of tumor-bearing, randomly selected mice. T2- and T2*-weighted MR
images acquired before and at different times after injection are shown in Figure
3.20. One can see that part of the tumor exhibits hypointensity 24 hours after the
injection of the conjugates; hence, a successful bond of magnetic nanocrystals to the
tumor via rch 24 mAb was established. More detailed evolvement with time has
been studied.
The T2 values gradually decreased and then reached a maximum of approximately 10% 24 hours after injection. In principle, these results suggest that the signal decrease should be caused mainly by the enrichment of the magnetic nanocrystals on the cancer cells mediated by rch 24 mAb. One can find the inhomogeneous distribution of the Fe3O4 nanocrystals, which was observed by Huh et al.
[153]. Their work is worth mentioning. In 2005, they reported an in vivo diagnosis
of cancer, with employment of a well-defined magnetic nanocrystal probe system
with multiple capabilities, such as small size, strong magnetism, high
biocompatibility, and the possession of active functionality for desired receptors.
The magnetic nanocrystals were conjugated to a cancer-targeting antibody, herceptin. The subsequent utilization of these conjugates as MRI probes was successfully
demonstrated for the monitoring in vivo of selective targeting events of human cancer cells implanted in live mice. Further conjugation of these nanocrystal probes
with fluorescent dyelabeled antibodies enables both in vitro and ex vivo optical
detection of cancer as well as in vivo MRI, which are potentially applicable for an
Before
24 h
Before
10 min
6h
10 h
24 h
(b)
(a)
(c)
Figure 3.20 (a) T2-weighted (upper row) and T2*-weighted (lower row) MR images acquired
before and after the injection of Fe3O4-(rch 24 mAb) conjugates. (b) T2*-weighted MR images
acquired before and at different times after injection of Fe3O4-(rch 24 mAb) conjugates (upper row)
and Fe3O4@PEG-COOH nanocrystals (lower row). (c) Histochemical analysis of a tumor tissue slice.
(Courtesy of Professor Mingyuan Gao, Institute of Chemistry, Chinese Academy of Science.)
82
advanced multimodal detection system. High-performance in vivo MR diagnosis of

cancer was achievable by utilizing improved and multifunctional material properties
of iron oxide nanocrystal bioprobes. In both groups the inhomogeneous distribution
of the magnetic nanocrystals was observed and was attributed to the heterogeneity
of the tumor.
In huge contrast, the PEG-coated Fe3O4 nanocrystals gave rise to nearly no variation either in the T2*-weighted images or T2 values of the tumor site over the entire
inspection time without the attachment of rch 24 mAb. Hence, the hypothesis that
the Fe3O4-(rch 24 mAb) conjugates can reach the tumor site and specifically bind
with the cancer cells LS 180 with CEA markers expressed on the cell membrane is
strongly supported. The researchers stated that the nude mice recovered from anesthesia spontaneously after the experiment and lived normally for weeks. This indicates that the bioconjugates have no acute fatal toxicity.
Tumor tissue slices were prepared and examined by successively staining with
an acid solution of potassium ferrocyanide, for developing ferric ions, and basic fuchsin
(4-[(4-aminophenyl)-(4-imino-3-methyl-1-cyclohexa-2,5-dienylidene)-methyl]aniline
hydrochloride), for cells.
In addition, novel carbon nanotubemetal cluster systems were advanced as
prototype systems for molecular recognition at the nanoscale. Ab initio calculations
indicate that the bare nanotube cluster system exhibits some specificity because the
adsorption of ammonia on a carbon nanotube (CNT)Al cluster system is easily
detected electrically, while diborane adsorption does not provide an electrical signature. Because there are well documented procedures for attaching molecular receptors to metal clusters, these results provide a proof of principle for the
development of novel, high-specificity molecular sensors. In 2005, Zhao et al. [149]
reported that nanotubemetal cluster systems were indeed promising systems for the
design of novel molecular sensors. Although the CNT-Al13 assembly is not reusable
for NH3 sensing because of the large adsorption energy, given the great flexibility in
functionalization of metallic clusters [150], the CNTmetal cluster assemblies could
be modified for recognition of specific chemical agents with high selectivity and
specificity. The key concepts are to use relatively small clusters and receptors that
donate or accept a significant amount of charge upon adsorption of a target molecule so that electron transport in the nanotube will be influenced. Charge transfer of
~0.1 electron per molecule is sufficient. From this fact we suggest that the metallic
NPs may be used together with C nanotubes for biopurposes.
As far as drug delivery is concerned, most research employs magnetite as the carrier or direction finder. In 2006, Cinteza et al. [160] reported the design, synthesis
utilizing nanochemistry, and characterization of a novel multifunctional polymeric
micelle-based nanocarrier system, where combined function of magnetophoretically
guided drug delivery together with light-activated photodynamic therapy was demonstrated. Specifically, the nanocarrier consists of polymeric micelles of
diacylphospholipid-poly-(ethylene glycol) (PE-PEG) coloaded with the photosensitizer drug 2-[1-hexyloxyethyl]-2-devinyl pyropheophorbide-a (HPPH), and
magnetic nanoparticles. The nanocarrier shows excellent stability and activity over
several weeks (Figure 3.21).
The physicochemical characterizations were carried out by means of TEM and
optical spectroscopy. An efficient cellular uptake has been confirmed with confocal
3.5 Specific Properties of Quantum Dots
83
Retention of HPPH, %
100
80
60
40
20
0
0
10
20
30
40
Time, hrs
50
60
70
Figure 3.21 The retention of HPPH in PEG-2000-PE magnetic polymeric micelles (10% magnetic
material) in PBS, pH = 7.4 at 37C. (From: [160]. 2006 American Chemical Society. Reprinted
with permission.)
laser scanning microscopy. They claimed that the loading efficiency of HPPH was
almost unaffected upon coloading with the magnetic nanoparticles, and its phototoxicity was retained. The magnetic response of the nanocarriers was demonstrated
by their magnetically directed delivery to tumor cells in vitro. The magnetophoretic
control on the cellular uptake provides enhanced imaging and phototoxicity.
Figure 3.22 indicates that the cells took up the HPPH entrapped in PEG-PE
micelles successfully. Localization of HPPH inside the cells [Figure 3.22(c)] is similar
to that of fluorescent micelles [Figure 3.22(b)], indicating that PEG-PE micelles keep
HPPH in cells after the treatment with a duration of 90 minutes. Hence, the similarity
of the distribution patterns obtained for fluorescence from fluorescein-labeled
micelles and those from encapsulated HPPH gives the opportunity to draw the conclusion that micellar nanocarriers sustained enough stability to retain HPPH during
incubation with cells in medium and after cellular uptake. In addition, the cells were
found to be in good condition even after staining overnight. In the same report, one
can see the remarkable achievement in magnetophoretic control and guidance of
cellular uptake by an external magnetic field [160].
3.5
Specific Properties of Quantum Dots

Research on quantum dots (QDs) has evolved over the past two decades from material science to biological applications. QDs, also known as nanocrystals, are a special class of semiconductors, which can be synthesized from various types of
semiconductor materials from periodic groups of II-VI (CdS, CdSe, CdTe, and so
forth), III-V (InP, InAs, and so forth), or IV-VI (PbSe and so forth). QDs are spherical nanocrystals, ranging from 1 to 10 nm (10 to 50 atoms) in diameter. Semiconductor nanocrystals can also be produced with other shapes such as rods and
tetrapods, but spherical QDs are the most widely used for biological applications.
QDs exhibit strongly size-dependent optical and electrical properties. A theoretical framework for these size-dependent properties was first described by Ekimov
84
(a)
(b)
(c)
Figure 3.22 Confocal microscopy images of Hela cells after 90 minutes of incubation with fluorescent PEG-2000-PE micelles loaded with HPPH: (a) transmission, (b) fluorescence from
fluorescein-labeled micelles, and (c) fluorescence from HPPH. (From: [160]. 2006 American
Chemical Society. Reprinted with permission.)
and Onushchenko [161] and Efros and Efros [162] in 1982. The physical properties
and applications of QDs have been extensively investigated in many physics and
engineering laboratories since the early 1980s [163165]. The size and shape can be
precisely controlled by duration, temperature, and the ligant molecules used in the
synthesis [166]. At these small sizes, materials behave differently, giving quantum
dots unprecedented properties and enabling never before seen applications in science and technology.
The optical and electronic qualities of traditional semiconductors are difficult to
adjust because their bandgap is fixed, and as a consequence they have fixed emission
frequencies. In contrast with conventional semiconductors, QDs have the ability to
tune the bandgap and hence the emission wavelength. The tunable bandgap of QDs
derives from the physical dimension of QDs, which by definition is smaller than the
exciton Bohr radius. In this case, the electron energy levels must be treated as discrete, and this gives rise to a unique phenomenon known as quantum confinement.
Quantum confinement, which refers to the spatial confinement of charge carriers
(i.e., electrons and holes) within a material, gives QDs unique optical and electronic
properties not observed in larger crystallites. With QDs, the size of the bandgap is
controlled simply by adjusting the size of the dot, and therefore it is possible to control the output wavelength of a dot with extreme precision. They exhibit a sizedependent color, as shown in Figure 3.23(a). However, it has been demonstrated
that the size differences of multicolor QDs could be addressed by using alloyed semiconductor QDs, where tuning of the emission wavelength is achieved via nanocrystal composition rather than size [Figure 3.23(b)].
QDs display advantages in their absorptive properties too, since they display
tunable absorption patterns. The absorption spectrum for quantum dots appears as
a series of overlapping peaks that get larger at shorter wavelengths. The dots will
absorb light that has a wavelength shorter than that of the first exciton peak, also
referred to as the absorption onset. The wavelength of the first exciton peak (and all
subsequent peaks) is a function of the composition and size of the dot.
In QDs the emission pattern is also tunable. The peak emission wavelength has a
Gaussian shape and occurs at a slightly longer wavelength than the absorption
3.5 Specific Properties of Quantum Dots
85
onset. This energy separation is what is referred to as the Stokes shift. An interesting
characteristics of QDs is that the peak emission wavelength is independent of the
wavelength of the excitation light, assuming that it is shorter that the wavelength of
the absorption onset. Many sizes of nanocrystals may therefore be excited with a
single wavelength of light, resulting in many colors that may be detected simultaneously. The absorption onset and the emission maximum shift to higher energy
with decreasing size of nanocrystals [166]. By varying the material used and different sizes for the nanocrystals, a spectral range from 400 nm to 2 m for emission
was obtained [167170]. The bandwidth of the emission spectra denoted as the full
width at half maximum (FWHM) has typical values of 20 to 30 ns in the visible
region of the spectrum and large extinction coefficients in the visible and ultraviolet
range. Narrower size distributions yield smaller FWHM. For CdSe a 5% size distribution corresponds to ~30-nm FWHM while in PbSe a 5% size distribution corresponds to ~100-nm FWHM [171].
The ratio of absorbed and emitted photons is called quantum yield (QY). QY is
controlled by the existence of nonradiative transitions of electrons and holes
between energy levelstransitions that produce no electromagnetic radiation.
Nonradiative recombination largely occurs at the dots surface and is therefore
greatly influenced by the surface chemistry. Passivation is the chemical process by
which the surface atoms of a QD are bonded to another material of much larger
bandgap. These (core) shell structures achieved efficient surface passivation as well
as carrier confinement within the QD core, which decreased the likelihood of
recombination via nonradiative pathways involving surface electronic states. In this
way QDs are coated with several atomic layers of an inorganic wideband semiconductor, and this results in a significant increase in the QY [172174]. The shell can
be designed carefully to obtain a QY close to 90% [175]. By bonding appropriate
molecules to this shell surface like thiol, amine, nitrile, phosphonic acid, carboxylic
acid, the QDs can be dispersed or dissolved in nearly any solvent or incorporated
into a variety of inorganic and organic films. In addition to the surface chemistry
Quantum dot composition
2.1 nm
400 450
3.2 nm
7.5 nm
500 550 600 650

Wavelength (nm)
(a)
Fluorescence (a.u.)
Fluorescence (a.u.)
Quantum dot size
700
750
CdSe
550 600
CdTe
CdSexTe1-x
650 700 750 800 850

Wavelength (nm)
900
(b)
Figure 3.23 QD emission wavelength, tunable by changing the nanoparticle size and composition. (a) The emission spectrum of a CdSe QD may be adjusted within the visible spectrum (450 to
650 nm) by selecting the QD size between 2 and 7.5 nm. The relative sizes of these particles are
shown schematically below the fluorescence spectrum. (b) While keeping the QD size constant
(5-nm diameter) and varying the composition of the ternary alloy CdSexTex, the maximum emission wavelength may be tuned between 610 and 800 nm. (From: [170]. 2004 Elsevier. Reprinted
with permission.)
86
molecular coupling can be used to effectively alter the properties of the quantum dot
including brightness and electronic lifetime. Of the semiconductor families investigated to date, IIVI materials have shown the most promise, and consequently their
use in biological applications has predominated [170].
3.6
Quantum Dots as Fluorescent Biological Labels

Living organisms are built of cells that have typically 10-m dimensions. The cell
parts and the proteins inside the cells are much smaller, with a typical size of 5 nm,
which is comparable with the dimensions of QDs. The size comparison shows the
possibility of investigating cellular components without introducing much disturbance [176]. Understanding biological processes on the nanoscale level is a strong
driving force behind development of nanotechnology [170, 177]. In 1998, two
groups, one led by A. P. Alivisatos at the University of California, Berkeley, and
another led by S. Nie (then at Indiana University Bloomington), simultaneously
demonstrated that semiconductor QDs could be made water soluble and could be
conjugated with biological molecules [178, 179]. Now they have entered a commercial exploration period [171, 177, 180, 181].
The fact that nanoparticles exist in the same size domain as proteins makes QDs
suitable for biotagging or biolabeling. In order to interact with biological targets, a
biological or molecular coating acting as a bioinorganic interface should be attached
to the nanoparticle. Examples of biological coatings may include antibodies or
biopolymers such as collagen [182] that make the nanoparticles biocompatible.
QDs can be made water soluble for use in life science applications by coating the
quantum dot surface with PEG lipids. These lipid-coated QDs can be used by themselves for applications such as cell tracking or they can be functionalized with a carboxyl or amine linker group for subsequent conjugation to other biomolecules
[171]. A schematic representation of the structure of a QD is shown in Figure 3.24.
3.6.1
Disadvantages of Organic Dyes, Traditional Biological Labels
Traditionally, the favored materials used to label microscopic structures have been
organic dyes, which can be chemically engineered to adhere to a diverse variety of
cellular structures. After the dye comes into contact with the proper cellular structures, they become fluorescent if excited with a certain wavelength of light. The
emitted radiation has a peak wavelength controlled by the chemical nature of the
organic dye being used. Unfortunately, many disadvantages exist with this technique, most of them a result of the extremely limited absorptive and emissive capabilities of organic dyes. For example, the peak emission of organic dyes is fixed, with
each dye having a different preset emission wavelength (color) that is set by its
nature. Therefore, applications that make use of light frequencies that do not correspond to the emission peaks of preexisting organic dyes cannot be performed.
Another disadvantage is the narrow absorption spectrum of organic dyes. Dyes have
uneven absorption and emission peaks. The shape of their emission and absorption
peaks shows shoulders, which is a major disadvantage in applications that require
a Gaussian-type emission spectrum. They tend to display absorption peaks that are
3.6 Quantum Dots as Fluorescent Biological Labels
87
Biological coating
Core
Figure 3.24
Shell
Schematic representation of a QD structure.
not always in convenient regions of the spectrum, making the excitation of various
organic dyes challenging and costly. The last disadvantage is that of stability. The
lifetime of organic dyes varies but is generally low relative to that of other tagging
mechanisms.
The fact that cell autofluorescence is in the visible spectrum and can mask signals from labeled molecules with dyes and the requirement of long observation
times have created the need to look for other fluorescent biological labels. The QDs
are nanometer-sized semiconductor nanocrystals with superior fluorescent properties. In comparison with organic dyes, which have fixed emissions, QDs can be
made to emit light at any wavelength in the visible and infrared ranges and can be
inserted almost anywhere, including liquid solution, dyes, paints, epoxies, and
sol-gels. QDs can be attached to a variety of surface ligands and inserted into a variety of organisms for in vivo research. They have unique technical capabilities to
enable all new standards for fluorescent tagging, eliminating the shortcomings of
dyes.
3.6.2
Beneficial Quantum Dot Optical and Spectral Properties
QDs have distinctive optical and spectral properties that provide unique properties
and benefits for a rich variety of biomedical life science applications. Bruchez et al.
[178] and Chan and Nie [179] and their coworkers first highlighted the great promise of QDs in biological applications:
Higher brightness As biological probes, QDs are extremely bright, with large
quantum yields. Studies comparing the brightness of single CdSe/ZnS
core/shell QDs to that of single rhodamine 6G molecules indicate that the fluorescent photon flux in QDs is tenfold to a hundredfold greater.
Broadband absorption. In contrast to organic fluorophores that have narrow
absorption spectra, QDs have large absorption cross sections over a wide
bandwidth. They display the unique property that all light that has a shorter
wavelength than the emission wavelength can be absorbed, with increasing
strength at shorter wavelengths. Therefore, the peak emission wavelength is
88
effectively independent of the excitation source. The effective Stokes shift can
be tens to hundreds of nanometers. In the visible range of the spectrum, the
peak emission wavelength is shifted from the absorption onset by 15 nm.
Tunable and narrow emission bandwidth. QDs have peak emission wavelengths dependent upon their composition and size. Currently, peak emission
wavelengths are available from 490 to 620 nm in 20-nm increments. Peak
emission wavelengths from 350 to 480, 640 to 720, and 800 to 2,300 nm have
been demonstrated [171]. QDs emit light within a narrow Gaussian spectrum
without any shoulders and centered at the peak wavelength.
Higher resistance to photo bleaching. QDs are based on inorganic particles
that are inherently more photostable than organic molecules, and as such they
can survive orders of magnitude longer than organic fluorescent dyes under
intense illumination.
Long fluorescence lifetime. The fluorescent lifetimes (electronic lifetime) of
visibly emitting quantum dots have been measured to be 15 to 20 ns, independent of the emission wavelength. The fluorescence lifetime is orders of
magnitude longer than typical autofluoresence lifetimes and many multiples
of typical organic dye lifetimes.
Large two-photon absorption. QDs have large two-photon absorption cross
sections that allow for narrowband visible light to be emitted when
long-wavelength IR lasers are focused on the quantum dots.
Improved detection sensitivity. The optical properties of QDs lead to
improved detection sensitivity and simplification in experiment and instrument design. Due to their inorganic semiconductor-based composition, they
can be also be visualized with electron microscopes.
A comparison between absorption and fluorescence spectra of an organic dye

(fluorescein isothiocyanate, FITC) and a CdSe QD with identical emission wavelengths is illustrated in Figure 3.25.
3.7
Quantum Dots in Biomedical Applications

During the past few years, QDs have been tested in most biotechnological applications that use fluorescence, including DNA array technology, immunofluorescence
assays, and cell and animal biology. The following paragraphs serve to highlight
some of the recent developments in the area of biological labeling and imaging.
The benefits of using QD-based labels in immunoassays were realized by
exploiting the spectral multiplexing capability of these nanocrystal fluorophores,
that is, the ability to excite and detect several labeled species simultaneously using a
single light source [183, 184]. Spectral multiplexing can easily be incorporated into
spatially multiplexed assays such as microarrays and the like.
In the area of biosensors, QDs are particularly attractive due to their long-term
photostability, allowing real-time and continuous monitoring. QDs can be used in
sensing applications via FRET, whereby nonradiative energy transfer occurs
between the QD (donor) and an acceptor molecule [185, 186]. QDs have been
shown to be very efficient FRET donors with organic fluorophores, due to the large
350
89
Absorbance of fluorescence (a.u.)
Absorbance of fluorescence (a.u.)
3.7 Quantum Dots in Biomedical Applications
FITC
400
450
500
550 600
Wavelength (nm)
650
350
CdSe quantum dot
400
450
500
550
Wavelength (nm)
600
650
Figure 3.25 Absorption and fluorescence emission spectra of an organic dye (fluorescein
isothiocyanate, FITC, left) and a CdSe QD (right) with identical emission wavelengths. The relative
sizes of the dye and nanoparticle are also shown. (From: [170]. 2004 Elsevier. Reprinted with
permission.)
overlap between the quantum dot emission wavelength and the absorption spectra
of the dyes. Because the emission characteristics of the QDs can be continuously
tuned, it is possible to create a FRET donor for any number of organic dyes that
emit between approximately 510 and 640 nm [171, 187]. QDs can act as highly efficient donors with other acceptors as other QDs and metallic nanoparticles [188].
The high photostability of QDs relative to organic dyes allows for long-term
tracking of biological processes. Due to the unprecedented photostability and color
multiplexing properties, continuous observations using fluorescence and confocal
microscopes of multiple cellular components and cellular metabolism/transport can
be made over a period of hours to days [189, 190]. An impressive demonstration of
this capability was illustrated by microinjecting QD bioconjugates into Xenopus
embryos and monitoring their partitioning into various cells during tadpole development over a period of several days [191].
The long-term stability and brightness of QDs make them ideal candidates for
live cell and animal targeting and imaging.
Water-stabilized quantum dots have been introduced into large multicellular
animals (mice) to preferentially stain vascular and lymphatic systems tumors, showing that higher contrast and imaging depth can be obtained at a lower excitation
power than with organic dyes. In [192], imaging of targeted QD delivery in live
mice was achieved by intravenously injecting them with QDs functionalized with
antibodies to prostate-specific membrane antigen. Many studies have shown the
great potential of using quantum dots as new probes in vivo [193, 194].
QDs have been shown to operate well in Western blot assays and can serve to
replace the radioactive labels and expensive detection equipment that are currently
in use. Conjugated to secondary antibodies, they have been successful applied to
protein electrophoretic gels that were prepared with primary antibodies. QD-based
Western blot technology was used for ultrasensitive detection of tracer proteins
[195].
QD fluor dyes are ideally suited for flow cytometry applications as recorders
and standards. For example, QDs 525 and 605 are useful for analyzing antigenic
expression. Flow cytometry (FCM), which is well adapted to detect fluorescence
90
emission of QDs in the UV or visible excitation mode, confocal laser scanning

microscopy (CLSM), and subsequent spectral analysis assess more specific characterization of QD fluorescent emissions [196]. The number of emission colors made
possible by quantum dots as well as the possibility of eliminating multiple lasers
sources of conventional flow cytometers and replacing them with simplified LEDs
and optics greatly increases throughput and will reduce the costs of next-generation
flow cytometer systems.
Two-photon excitation fluorescence cross-correlation spectroscopy (TPE-XCS)
is a very suitable method for studying interactions of two distinctly labeled fluorescent molecules. When standard organic fluorescent labels are used, there is almost
inevitably spectral cross talk between the detection channels, which must be
accounted for in TPE-XCS data analysis. However, using quantum dots as labels for
both ligand and receptor, this limitation can be alleviated, because of the dots narrower emission spectra [197]. Advantages include the ability to greatly reduce
autofluorescence of the sample as well as the ability to use a greater range of excitation sources that are not absorbed by the sample.
The long fluorescence lifetime of QDs (15 to 20 ns) enables the use of time-gated
detection [198] to separate their signals from those of shorter lived species, such as
background autofluorescence encountered in cells. When the fluorescence lifetime
instrumentation currently available is used, greater signal-to-noise ratios are
possible.
Following the first simultaneous reports of using QDs as labels in biological
experiments by Bruchez et al. [178] and Chan and Nie [179], the number of QD biological studies has increased exponentially. The breakthroughs and steady improvements made over the past 10 years in producing and modifying QDs for use as
fluorescent labels have contributed to the successful implementation of this new and
unique probe. Although they are unlikely to completely replace traditional organic
fluorophores as biological labels, QDs have secured their place as a viable technology in the biological sciences. With their capability for single-molecule and multiplexed detection, real-time imaging, and biological compatibility, QDs represent a
valuable new technology offering numerous benefits that can enable researchers and
product developers to create new assays or even new diagnostics.
Besides their superior properties as fluorescent labels, the increasingly popular
application of QD-based labels for biological studies of living organisms has
required that QD toxicity be examined since it is well known that QDs contain toxic
elements. Studies have shown that quantum dot toxicity is due to cadmium ions
being released into cells. This is exacerbated by oxygen, UV exposure, and the large
exposed surfaces of the spherical quantum dots. To avoid these problems, surfaces
are added to the quantum dots, traditionally consisting of zinc-sulfide compounds.
Polymers are used as well, which have been shown to be stable, but the polymers
themselves may kill cells. Studies of quantum dot cytotoxicity are available in
[199201]. In 2006, Evident Technologies announced that they had made QDs that
contain no heavy metals commercially available [171].
Recent developments in QD synthesis and bioconjugation, their applications in
molecular and cellular imaging, as well as promising directions for future research
are reviewed in [170, 177, 202208].
91
References
[1] Trindade, T., P. OBrien, and N. L. Pickett, Nanocrystalline Semiconductors: Synthesis,
Properties, and Perspectives, Chem. Mater., Vol. 13, 2001, p. 3843.
[2] Burda, C., et al., Chemistry and Properties of Nanocrystals of Different Shapes, Chem.
Rev., Vol. 105, 2005, p. 1025.
[3] Bohren, C. F., and D. R. Huffman, Absorption and Scattering of Light by Small Particles,
New York: Wiley, 1983.
[4] Creighton, J. A., and D. G. Eadon, New Mixed Quantum/Semiclassical Propagation
Method, J. Chem. Soc. Faraday Trans., Vol. 87, 1991, p. 3881.
[5] Ostwald, W., Theoretical and Applied Colloid Chemisty, New York: Wiley, 1922, p. 200.
[6] Kreibig, U., and L. Genzel, Optical Absorption of Small Metallic Particles, Surf. Sci., Vol.
156, 1985, p. 678.
[7] Fragstein, C. V., and U. Z. Kreibig, Studies of Intersystem Crossing Dynamics in Acetylene, Phys., Vol. 224, 1969, p. 306.
[8] Doremus, R. H., Optical Properties of Small Silver Particle, J. Chem. Phys., Vol. 42,
1965, p. 414.
[9] Doyle, W. T., Absorption of Light by Colloids in Alkali Halide Crystals, Phys. Rev., Vol.
B111, 1958, p. 1067.
[10] Huffman, D. R., in An Introduction to Physiscs and Technology of Thin Films,
Wagendristerl, A., and Wang, Y., (eds.), New Jersey: World Scientific, 1988.
[11] Ralph, D. C., C. T. Black, and M. Tinkham, Spectroscopy of the Superconducting Gap in
Individual Nanometer-Scale Aluminum Particles, M. Phys. Rev. Lett., Vol. 74, 1995, pp.
32413244.
[12] Peng, X., J. Wickham, and A. P. Alivisatos, Kinetics of II-VI and III-V Colloidal Semiconductor Nanocrystal Growth: Focusing of Size Distributions, J. Am. Chem. Soc., Vol.
120, 1998, p. 5343.
[13] Murray, C. B., C. R. Kagan, and M. G. Bawendi, The Width of the Absorption Band,
Annu. Rev. Mater. Sci.,Vol. 30, 2000, p. 545.
[14] Pamplin, B. R., Crystal Growth, New York: Pergamon Press, 1975.
[15] Jiang, Y., Forced Hydrolysis and Chemical Co-Precipitation, in Handbook of
Nanophase and Nanostructured Materials, Z. L.Wang, Y. Liu, and Z. Zhang, (eds.), New
York: Kluwer Academic, 2003, p. 59.
[16] Burda, C., et al., Chemistry and Properties of Nanocrystals of Different Shapes, Chem.
Rev., Vol. 105, 2005, p. 1025.
[17] Alivisatos, A. P., Naturally Aliened Nanoparticles, Science, Vol. 289, 2000, p. 736.
[18] Hayat, M. A., (ed.), Colloidal Gold: Principles, Methods and Applications, San Diego, CA:
Academic Press, Vols. 1 and 2, 1989.
[19] Brust, M., et al., Ultrafast Optical Kerr Effect Spectroscopy of Water Confined in
Nanopores of the Gelatin Gel, Chem. Soc. Chem. Commun., 1994, p. 801.
[20] Johnson, S. R., et al., Alkanethiol Molecules Containing an Aromatic Moiety Self-Assembled onto Gold Clusters, Langmuir, Vol. 13, 1997, p. 51.
[21] Hostetler, M. J., et al., Monolayers in Three Dimensions: Synthesis and Electrochemistry
of Functionalized Alkanethiolate-Stabilized Gold Cluster Compounds, Am. Chem. Soc.,
Vol. 118, 1996, p. 4212.
[22] Ingram, R. S., M. J. Hostetler, and R. W. J. Murray, Poly-HeteroFunctionalized
Alkanethiolate-Stabilized Gold Cluster Compounds, Am. Chem. Soc., Vol. 119, 1997, p.
9175.
[23] Buining, P. A., et al., Preparation of Functional Silane-Stabilized Gold Colloids in the
(Sub)nanometer Size Range, Langmuir, Vol. 13, 1997, p. 3921.
92

[24] Brown, L. O., and J. E. Hutchison, Convenient Preparation of Stable, Narrow-Dispersity,
Gold Nanocrystals by Ligand Exchange Reactions, J. Am. Chem. Soc., Vol. 119, 1997, p.
12384.
[25] Brust, M., et al., Influence of Vapor Depletion on Nucleation Rate, J. Chem. Soc. P.
Chem. Commun. 1995, pp. 16551656.
[26] Yonezawa, T., M. Sutoh, and T. Kunitake, Ab Initio Melting Curve of Molybdenum by the
Phase Coexistence Method, Chem. Lett., Vol. 619, 1997, p. 2365.
[27] Chen, S., and K. Kimura, Synthesis and Characterization of Carboxylate-modified Gold
Nanoparticle Powders Dispersible in Water Langmuir, Vol. 15, 1999, pp. 10751082.
[28] Leff, D. V., et al, Thermodynamic Control of Gold Nanocrystal Size: Experiment and Theory,J. Phys. Chem., Vol. 99, 1995, pp. 70367041.
[29] Brust, M., et al., Columnar Versus Smectic Order in Systems of Charged Boble
Nanoparticles, J. Chem. Soc. Chem. Commun., 1995, pp. 16551656.
[30] Ralph, D. C., C. T. Black, and M. Tinkham, Spectroscopic Measurements of Discrete Electronic States in Single Metal Particles, Phys. Rev. Lett., Vol. 74, 1995, p. 3241.
[31] Huffman, D. R., in An Introduction to Physiscs and Technology of Thin Films, A.
Wagendristerl and Y. Wang, (eds.), New York: World Scientific, 1988.
[32] Dubois, L. H., and R. G. Nuzzo, Nonlocal Optical Effects on the Fluorescence and Decay
Rates for Admolecules at a Metallic Nanoparticle, Annu. Rev. Phys. Chem., Vol. 43,
1992, p. 437.
[33] Fenter, P., A. Eberhardt, and P. Eisenberger, Self-Assembly of n-Alkyl Thiols as Disulfides
on Au(111), Science, Vol. 266, 1994, p. 1216.
[34] He, S. J., et al., Formation of Silver Nanoparticles and Self-Assembled Two-Dimensional
Ordered Superlattice, Langmuir, Vol. 17, 2001, p. 1571.
[35] Zhang, Z., et al., Stable Silver Clusters and Nanoparticles Prepared in Polyacrylate and
Inverse Micellar Solutions, J. Phys. Chem. B, Vol. 104, 2000, p. 1176.
[36] Peng, X., J. Wickham, and A. P. Alivisatos, Kinetics of II-VI and III-V Colloidal Semiconductor Nanocrystal Growth: Focusing on Size Distribution, J. Am. Chem. Soc., Vol. 120,
1998, p. 5343.
[37] Valden, M., X. Lai, and D. W. Goodman, Onset of Catalytic Activity of Gold Clusters on
Titania with the Appearance of Nonmetallic Properties, Science, Vol. 281, 1998, p. 1647.
[38] Link, S., and M. A. El-Sayed, Spectral Properties and Relaxation Dynamics of Surface
Plasmon Electronic Oscillations in Gold and Silver Nanodots and Nanorods, J. Phys.
Chem. B, Vol. 103, 1999, p. 8410.
[39] He, S. T., et al., Self-Assembled Two-Dimensional Superlattice of AuAg Alloy Nanocrystals, Appl. Phys. Lett., Vol. 81, 2002, p. 150.
[40] Wang, Z. L., Transport and Selectivity in Nanopores with Spatially Inhomogeneous
Nanoparticles, Aust. J. Chem., Vol. 54, 2001, p. 153.
[41] Petit, C., A. Talab, and M. P. Pileni, Cobalt Nanosized Particles Organized in a 2D
Superlattice: Synthesis, Characterization, and Magnetic Properties, J. Phys. Chem. B, Vol.
103, 1999, p. 1805.
[42] Gao, H. J., et al., Reversible, Nanometer-Scale Conductance Transitions in an Organic
Complex, Phys. Rev. Lett., Vol. 84, 2000, p. 1780.
[43] Giersig, M., and P. Mulvaney, Preparation of Ordered Colloid Monolayers by Electrophoretic Deposition, Langmuir, Vol. 9, 1993, p. 3408.
[44] Feldstein, M. J., et al., Electronic Relaxation Dynamics in Coupled Metal Nanoparticles,
J. Am. Chem. Soc., Vol. 119, 1997, p. 6638.
[45] Puntes, V. F., K. M. Krishan, and A. P. Alivisatos, Colloidal Nanocrystal Shape and Size
Control: The Case of Cobalt, Science, Vol. 291, 2001, p. 2115.
[46] Murray, C. B., D. J. Norris, and M. G. Bawendi, Synthesis and Characterization of Nearly
Monodisperse Cde (E = Sulfur, Selenium, Tellurium) Semiconductor Nanocrystallites, J.
Am. Chem. Soc., Vol. 115, 1993, p. 8706.
93
[47] Kitakami, O., et al., Size Effect on the Crystal Phase of Cobalt Fine Particles, Phys. Rev.
B, Vol. 56, 1997, p. 13849.
[48] Hull, A. W., X-Ray Crystal Analysis of Thirteen Common Metals, Phys. Rev., Vol. 17,
1921, p. 571.
[49] Krainer, E., and J. Robitsch, New Diffraction in Special Bulk Cobalt Surfaces, 2
Metal//kd, Vol. 65, 1974, p. 729.
[50] Kajivara, S., et al., Computational Study of the Melting-Freezing Transition in the Quantum Hard-Sphere System, Phil. Mag. Lett., Vol. 55, 1987, p. 215.
[51] Leslie-Pelecky, D. L., et al., Using High-Temperature Chemical Synthesis to Produce
Metastable Nanostructured Cobalt, Chem. Mater., Vol. 10, 1998, p. 3732.
[52] Respaud, M., et al., Surface Effects on the Magnetic Properties of Ultrafine Cobalt Particles, Phys. Rev. B, Vol. 57, 1998, p. 2925.
[53] Dinega, D. P., and M. G. Bawendi, A Solution-Phase Chemical Approach to a New Crystal Structure of Cobalt, Angew. Chem. Int. Ed., Vol. 38, No. 12, 1999, pp. 17881791.
[54] Sun, S., and C. B. Murray, Synthesis of Monodisperse Cobalt Nanocrystals and Their
Assembly into Magnetic Superlattices, J. Appl. Phys., Vol. 85, 1999, p. 4325.
[55] Gomez, S., et al., Water-Gas-Shift Reaction on Metal Nanoparticles and Surfaces, Chem.
Commun., 2001, p. 1474.
[56] Gomez, S., et al., Theoretical Study of the Electronic Structure of Silver Nanoparticles,
Chem. Commun., 2000, p. 1945.
[57] Dinega, D. P., and M. G. Bawendi, A Solution-Phase Chemical Approach to a New Crystal Structure of Cobalt, Angew. Chem. Int. Ed., Vol. 38, 1999, p. 1788.
[58] Zhang, P., and T. K. Sham, Tuning the Electronic Behavior of Au Nanoparticles with Capping Molecules, Appl. Phys. Lett., Vol. 81, 2002, p. 736.
[59] Selwood, P. W., Chemisorption and Magnetization, New York: Academic Press, 1975.
[60] Puntes, V. F., et al., Synthesis of hcp-Co Nanodisks, J. Am. Chem. Soc., Vol. 124, 2002,
p. 12874.
[61] Bradley, J. S., et al., Surface Chemistry on Colloidal Metals: Spectroscopic Study of
Adsorption of Small Molecules, Faraday Discuss., Vol. 92, 1991, p. 255.
[62] Pan, C., et al., Ligand-Stabilized Ruthenium Nanoparticles: Synthesis, Organization, and
Dynamics, J. Am. Chem. Soc., Vol. 123, 2001, p. 7584.
[63] Choukroun, R., et al., H2-Induced Structural Evolution in Non-Crystalline Rhodium
Nanoparticles, New J. Chem., Vol. 25, 2001, p. 525.
[64] Rodriguez, A., et al., Synthesis and Isolation of Cuboctahedral and Icosahedral Platinum
Nanoparticles. Ligand-Dependent Structures, Chem. Mater., Vol. 8, 1996, p. 1978.
[65] Osuna, J., et al., Chemical Control of Structural and Magnetic Properties of Cobalt
Nanoparticles, Chem. Mater., Vol. 17, 2005, p. 107.
[66] Pery, T., et al., Direct NMR Evidence for the Presence of Mobile Surface Hydrides on
Ruthenium Nanoparticles, Angew. Chem., Int. Ed., Vol. 6, 2005, p. 605.
[67] Cordente, N., et al., Synthesis and Magnetic Properties of Nickel Nanorods, Nano Lett.,
Vol. 1, 2001, p. 565.
[68] Bnnemann, H., et al., A Size-Selective Synthesis of Air Stable Colloidal Magnetic Cobalt
Nanoparticles, Inorg. Chim. Acta, Vol. 350, 2003, p. 617.
[69] Dassenoy, F., et al., Experimental Evidence of Structural Evolution in Ultrafine Cobalt
Particles Stabilized in Different PolymersFrom a Polytetrahedral Arrangement to the
Hexagonal Structure, J. Chem. Phys., Vol. 112, 2000, p. 8137.
[70] Petit, C., A. Taleb, and M.-P. Pileni, Self-Organization of Magnetic Nanosized Cobalt
Particles, Adv. Mater., Vol. 10, 1998, p. 259.
[71] Andersen, R. A., et al., Synthesis of bis[bis(trimethylsilyl)amido]iron(II). Structure and
Bonding in M[N(SiMe3)2]2 (M=Manganese, Iron, Cobalt): Two-Coordinate Transition-Metal Amides, Inorg. Chem., Vol. 27, 1988, p. 1782.
94

[72] Hull, A. W., X-Ray Crystal Analysis of Thirteen Common Metals, Phys. Rev., Vol. 17,
1921, p. 571..
[73] Krainer, E., Selective Spin Diffusion. A Novel Method for Studying Motional Properties of
Biopolymers in Solution, Metallkd, Vol. 65, 1974, p. 729.
[74] Respaud, M., et al., Surface Effects on the Magnetic Properties of Ultrafine Cobalt Particles, Phys. Rev. B, Vol. 57, 1998, p. 2925.
[75] Shoemaker, C. B., et al., Refinement of the Structure of Manganese and of a Related Phase
in the Mn-Ni-Si System, Acta Crystallogr. Sect. B, Vol. 34, 1978, p. 3573.
[76] Landolt-Brnstein, in Numerical Data and Functional Relationships in Science and Technology, Vol. 19/a, K. H. Hellwege and O. Madelung, (eds.), Berlin: Springer, 1986, p. 36.
Monodisperse CdE (E=Sulfur, Selenium, Tellurium) Semiconductor Nanocrystallites, J.
Am. Chem. Soc., Vol. 115, 1993, p. 8706.
[78] Yang, T., et al., Highly Ordered Self-Assembly with Large Area of Fe3O4 Nanoparticles
and the Magnetic Properties, J. Phys. Chem. B, Vol. 109, 2005, p. 23233.
[79] Yang, H. T., et al., Self-Assembly and Magnetic Properties of Cobalt Nanoparticles,
Appl. Phys. Lett., Vol. 82, 2003, p. 4729.
[80] Murray, C. B., et al., Growth Dynamics and Characterization of Magnetic Nanoparticles, IBM J. Res. Dev., Vol. 45, 2001, p. 47.
[81] Fauth, K., U. Kreibig, and G. Z. Schmid, Inert Gas Condensation of Evaporated Ni and
Laser Ablated Co, Phys. D., Vol. 12, 1989, p. 515.
[82] Duff, D. G., A. Baiker, and P. P. Edwards, A Transient Electrical Model of Charging for Ag
Nanocrystal, J. Chem. Soc. Chem. Commun., 1993, p. 96.
[83] Mulvaney, P., Surface Plasmon Spectroscopy of Nanosized Metal Particles, Langmuir,
Vol. 12, 1996, p. 788.
[84] Alvarez, M. M., et al., Optical Absorption Spectra of Nanocrystal Gold Molecules, J.
Phys. Chem. B, Vol. 101, No. 19, 1997, pp. 37063711.
[85] Terrill, R. H., Monolayers in Three Dimensions: NMR, SAXS, Thermal, and Electron
Hopping Studies of Alkanethiol Stabilized Gold Clusters, J. Am. Chem. Soc., Vol. 117,
1995, p. 12537.
[86] Badia, A., Structure and Chain Dynamics of Alkanethiol-Capped Gold Colloids,
Langmuir, Vol. 12, 1996, p. 1262.
[87] Harfenist, S. A., et al., Highly Oriented Molecular Ag Nanocrystal Arrays, J. Phys.
Chem., Vol. 100, 1996, p. 13904.
[88] Brust, M., et al., Novel Gold-Dithiol Nano-Networks with Non-Metallic Electronic Properties, Adv. Mater., Vol. 7, 1995, p. 795.
[89] Whetten, R. L., et al., Nanocrystal Gold Molecules, Adv. Mater., Vol. 8, 1995, p. 428.
[90] Luedtke, W. D., and U. Landman, Structure, Dynamics, and Thermodynamics of Passivated Gold Nanocrystallites and Their Assemblies, J. Phys. Chem., Vol. 100, 1996, p.
13323.
[91] Alvarez, M. M., et al., Critical Sizes in the Growth of Au Clusters, Chem. Phys. Lett.,
Vol. 266, 1997, p. 91.
[92] Duff, D. G., A. Baiker, and P. P. Edwards, Visualization of Induced Charge in a
Nanoparticles System, J. Chem. Soc., Chem. Commun., 1993, p. 96.
[93] Lee, H., et al., Antibiofouling Polymer-Coated Superparamagnetic Iron Oxide
Nanoparticles as Potential Magnetic Resonance Contrast Agents for In Vivo Cancer Imaging, J. Am. Chem. Soc., Vol. 117, 2006, p. 7383.
[94] Taylor, K. J., et al., Ultraviolet Photoelectron Spectra of Coinage Metal Clusters, J.
Chem. Phys., Vol. 96, 1992, p. 3319.
[95] Frankamp, B. L., A. K. Boal, and V. M. Rotello, Controlled Interparticle Spacing Through
Self-Assembly of Au Nanoparticles and Poly(amidoamine) Dendrimers, J. Am. Chem.
Soc., Vol. 124, 2002, p. 15146.
95
[96] Ostwald, W., Theoretical and Applied Colloid Chemisty, New York: Wiley, 1922, 200.
[97] Johnson, P. B., and R. W. Christy, Monodispersed Particles of Gold, Phys. Rev., 1972,
Vol. B6, 1972, pp. 43704379.
[98] Ziman, J. M., Theory of Solids, Cambridge, U.K.: Cambridge University Press, 1979, p.
278.
[99] Fang, M., et al., Magnetic Bio/Nanoreactor with Multilayer Shells of Glucose Oxidase
and Inorganic Nanoparticles, Langmuir, Vol. 18, 2002, p. 6338.
[100] Hovel, H. S., et al., Raman Scattering of 4-Aminobenzenethiol Sandwiched between
Ag/Au Nanoparticle and Macroscopically Smooth Au Substrate, Phys. Rev., Vol. B48,
1993, p. 18178.
[101] Henglein, A., Physicochemical Properties of Small Metal Particles in Solution:
Microelectrode Reactions, Chemisorption, Composite Metal Particles, and the
Atom-to-Metal Transition, J. Phys. Chem., Vol. 97, 1993, p. 5457.
[102] Camillone III, et al., Substrate Dependence of the Surface Structure and Chain Packing of
Docosyl Mercaptan Self-Assembled on the (111), (110), and (100) Faces of Single Crystal
Gold, J. Chem. Phys., Vol. 98, 1993, p. 4234.
[103] Linnert, T., P. Mulvaney, and A. J. Henglein, Surface Chemistry of Colloidal Silver: Surface Plasmon Damping by Chemisorbed Iodide, Hydrosulfide (SH-), and Phenylthiolate,
Phys. Chem., Vol. 97, 1993, p. 679.
[104] Kreibig, U., et al., in Large Clusters of Atoms and Molecules, T. P. Martin, (ed.), Dordrecht:
Kluwer, 1996, p. 477.
[105] Cleveland, C. L., et al., Structural Evolution of Larger Gold Clusters, Phys. D, Vol. 47,
1999, p. 468.
[106] Leslie-Pelecky, D. L., and R. D. Rieke, Magnetic Properties of Nanostructured Materials, Chem. Mater., Vol. 8, 1996, p. 1770.
[107] Selwood, P. W., Chemisorption and Magnetization, New York: Academic Press, 1975.
[108] Ruitenbeek, J. M. V., D. A. V. Leeuven, and L. J. D. Jongh, Magnetic Properties of Metal
Cluster Compounds, in Physics and Chemistry of Metal Cluster Compounds, L. J. D.
Jongh, (ed.), Dordrecht: Kluwer Academic, 1994, p. 277.
[109] Haes, A. J., et al., Detection of a Biomarker for Alzheimers Disease from Synthetic and
Clinical Samples Using a Nanoscale Optical Biosensor, J. Am. Chem. Soc., Vol. 127,
2005, p. 2264.
[110] Kohler, N., G. E. Fryxell, and M. Zhang, A Bifunctional Poly(ethylene glycol) Silane
Immobilized on Metallic Oxide-Based Nanoparticles for Conjugation with Cell Targeting
Agents, J. Am. Chem. Soc., Vol. 126, 2004, p. 7206.
[111] Hu, A., G. T. Yee, and W. U. Lin, Magnetically Recoverable Chiral Catalysts Immobilized
on Magnetite Nanoparticles for Asymmetric Hydrogenation of Aromatic Ketones, J. Am.
Chem. Soc., Vol. 127, 2005, p. 12486.
[112] Roucoux, A., J. Schulz, and H. Patin, Reduced Transition Metal Colloids: A Novel Family
of Reusable Catalysts? Chem. Rev., 2002, Vol. 102, p. 3757.
[113] Ingram, R. S., M. Hostetler, and R. W. Murray, Poly-HeteroFunctionalized
Alkanethiolate-Stabilized Gold Cluster Compounds, J. Am. Chem. Soc., Vol. 119, 1997,
p. 9175.
[114] Morrison, R. T., and R. N. Boyd, Organic Chemistry, 6th ed., Englewood Cliffs, NJ:
Prentice-Hall, 1992.
[115] Carey, F. A., and R. J. Sundberg, Advanced Organic Chemistry: Part A: Structure and
Mechanism, 3rd Ed., New York: Penum Press, 1990, p. 476.
[116] Templeton, A. C., et al., Gateway Reactions to Diverse, Polyfunctional MonolayerProtected Gold Clusters, J. Am. Chem. Soc., Vol. 120, 1998, p. 4845.
[117] Lwowski, W., in 1,3-Dipolar Cycloaddition Chemistry, A. Padwa (Ed.), New York: Wiley,
1984.
96

[118] Huisgen, R., Hybridization and Enzymatic Extension of Au Nanoparticle-Bound
Oligonucleotides, Pure Appl. Chem., Vol. 61, 1989, p. 613.
[119] Fleming, D. A., C. J. Thode, and M. E. Williams, Triazole Cycloaddition as a General
Route for Functionalization of Au Nanoparticles, Chem. Mater., Vol. 18, No. 9, 2006,
pp. 23272334.
[120] Brust, M., et al., Raman Scattering of 4-Aminobenzenethiol Sandwiched Between Ag/Au
Nanoparticle and Macroscopically Smooth Au Substrate, Chem. Soc. Chem. Commun.,
1994, p. 801.
[121] Templeton, A. C., et al., Reactivity of Monolayer-Protected Gold Cluster Molecules: Steric
Effects, J. Am. Chem. Soc., Vol. 120, 1998, p. 1906.
[122] Hostetler, M. J., A. C. Templeton, and R. W. Murray, Dynamics of Place-Exchange Reactions on Monolayer-Protected Gold Cluster Molecules, Langmuir, Vol. 15, 1999, p. 3782.
[123] Collman, J. P., N. K. Devaraj, and C. E. D. Chidsey, Clicking Functionality onto Electrode Surfaces, Langmuir, Vol. 20, 2004, p. 1051.
[124] Rispers, T., and J. B. F. N. Engberts, Growth of Gold Nanoparticles in Human Cells, J.
Phys. Org. Chem., Vol. 18, 2005, p. 908.
[125] Gajewski, J., The Claisen Rearrangement. Response to Solvents and Substituents: The
Case for Both Hydrophobic and Hydrogen Bond Acceleration in Water and for a Variable
Transition State, J. Acc. Chem. Res., Vol. 30, 1997, p. 219.
[126] Kolb, H. C., M. G. Finn, and K. B. Sharpless, Click Chemistry: Diverse Chemical Function
from a Few Good Reactions, Angew. Chem. Int., Vol. 40, 2001, p. 2004.
[127] Rostovtsev, V. V., et al., A Stepwise Huisgen Cycloaddition Process: Copper(I)-Catalyzed
Regioselective Ligation of Azides and Terminal Alkynes, Angew. Chem. Int. Ed., Vol. 41,
2002, p. 2596.
[128] Wu, P., et al., Efficiency and Fidelity in a Click-Chemistry Route to Triazole Dendrimers
by the Copper(I)-Catalyzed Ligation of Azides and Alkynes, Angew. Chem. Int. Ed., Vol.
43, 2004, p. 3928.
[129] Billing, J. F., and U. J. Nilsson, Noble Metal Clusters Encapsulated by Organozinc
Phosphazenate Assemblies, J. Org. Chem., Vol. 70, 2005, p. 4847.
[130] Van Maarseveen, J. H., W. S. Horne, and M. R. Ghadiri, Efficient Route to C2 Symmetric
Heterocyclic Backbone Modified Cyclic Peptides, Org. Lett., Vol. 7, 2005, p. 4503.
[131] Malinsky, M. D., et al., Chain Length Dependence and Sensing Capabilities of the Localized Surface Plasmon Resonance of Silver Nanoparticles Chemically Modified with
Alkanethiol Self-Assembled Monolayers, J. Am. Chem. Soc., Vol. 123, 2001, p. 1471.
[132] Haes, A. J., and R. P. Van Duyne, A Nanoscale Optical Biosensor: Sensitivity and Selectivity of an Approach Based on the Localized Surface Plasmon Resonance Spectroscopy of Triangular Silver Nanoparticles, J. Am. Chem. Soc., Vol. 124, 2002, p. 10596.
[133] Haes, A. J., and R. P. Van Duyne, Seeded Growth of Submicron Au Colloids with Quadrupole Plasmon Resonance Modes, Laser Focus World, Vol. 39, 2003, p. 153.
[134] Riboh, J. C., et al., A Nanoscale Optical Biosensor: Real-Time Immunoassay in Physiological Buffer Enabled by Improved Nanoparticle Adhesion, J. Phys. Chem. B, Vol. 107, 2003,
p. 1772.
[135] Haes, A. J., et al., A Nanoscale Optical Biosensor: The Long Range Distance Dependence
of the Localized Surface Plasmon Resonance of Noble Metal Nanoparticles, J. Phys.
Chem. B, Vol. 108, 2004, p. 109.
[136] Haynes, C. L., and R. P. Van Duyne, Nanosphere Lithography: A Versatile Nanofabrication Tool for Studies of Size-Dependent Nanoparticle Optics, J. Phys. Chem. B,
Vol. 105, 2001, p. 5599.
[137] Lee, J., Biocatalytic Evolution of a Biocatalyst Marker: Towards the Ultrasensitive Detection of Immunocomplexes and DNA Analysis, Angew. Chem. Int. Ed., Vol. 45, 2006, p.
4819.
97
[138] Braude, A. I., The Optical Properties of Metal Nanoparticles and the Bio Application, N.
Engl. J. Med., Vol. 258, 1958, p. 1289.
[139] Klein, H. G., et al., Complementary Bioconjugates Based on Antibody-Antigen with
Nanoparticles, J. Nemo, Transfusion, Vol. 37, 1997, p. 95.
[140] Lee, C. K., et al., Detectable Signal in the Luminescence Image in Bio-Nanoparticles, Vox
Sang., Vol. 82, 2002, p. 204.
[141] Goodnough, L. T., et al., Surfactantless Synthesis of Multiple Shapes of Gold
Nanostructures and Their Shape-Dependent SERS Spectroscopy, N. Engl. J. Med., Vol.
340, 1999, p. 438.
[142] Puckett, A., et al., Capacitance Measurements of Antibody-Antigen Interactions in a Flow
System, J. Clin. Pathol., Vol. 45, 1992, p. 155.
[143] McDonald, C. P., et al., Detection of a Biomarker for Alzheimers Disease from Synthetic
and Clinical Samples Using a Nanoscale Optical Biosensor, Transfus. Med., Vol. 6, 1996,
p. 61.
[144] Anderson, K. C., et al., Magnetic Resonance Imaging of the Brain in Williams-Beuren Syndrome, Am. J. Med., Vol. 81, 1986, p. 405.
[145] Gu, H. W., et al., Using Biofunctional Magnetic Nanoparticles to Capture VancomycinResistant Enterococci and Other Gram-Positive Bacteria at Ultralow Concentration, J.
Am. Chem. Soc., Vol. 125, 2003, No. 15, p. 702.
[146] Gu, H. W., et al., A Nanoscale Optical Biosensor Based on Localized Surface Plasmon
Resonance, Chem. Commun., 2003, p. 1966.
[147] Sun, S. H., et al., Monodisperse FePt Nanoparticles and Ferromagnetic FePt Nanocrystal
Superlattices, Science, Vol. 287, 2000, 1989.
[148] Performance Standards for Antimicrobial Susceptibility Testing, National Committee
for Clinical Laboratory Standards, Villanova, PA, 1998.
[149] Zhao, Q., et al., Carbon Nanotube-Metal Cluster Composites: A New Road to Chemical
Sensors, Nano. Lett., Vol. 5, 2005, p. 847.
[150] See, E. G., in Metal Nanoparticles: Synthesis, Characterization and Applications, C. A.
Foss, Jr. and D. L. Feldheim (Eds.), New York: Marcel Dekker, 2002.
[151] Hu, F., et al., Preparation of Biocompatible Magnetite Nanocrystals for In Vivo Magnetic
Resonance Detection of Cancer, Adv. Mater., Vol. 19, 2006, p. 2553.
[152] Such, C. C., A. S. Berry, and G. Curtis, Functionalisation of Magnetic Nanoparticles for
Applications in Biomedicine, J. Phys. D: Appl. Phys., Vol. 36, 2003, p. R198.
[153] Huh, Y.-M., et al., In Vivo Magnetic Resonance Detection of Cancer by Using Multifunctional Magnetic Nanocrystals, J. Am. Chem. Soc., Vol. 127, 2005, p. 12387.
[154] Zhao, M., et al., Silica-Encapsulated Nanomagnetic Particle as a New Recoverable Biocatalyst Carrier, Nat. Med., Vol. 7, 2001, p. 1241.
[155] Weissleder, R., et al., Long-Circulating Iron Oxides for MR Imaging, Adv. Drug Delivery Rev., Vol. 16, 1995, p. 321.
[156] Gupta, A. K., and M. Gupta, Synthesis and Surface Engineering of Iron Oxide
Nanoparticles for Biomedical Applications, Biomaterials, Vol. 26, 2005, p. 3995.
[157] Meldrum, F. C., B. R. Heywood, and S. Mann, Magnetoferritin: In Vitro Synthesis of a
Novel Magnetic Protein, Science, Vol. 257, 1992, p. 522.
[158] Bulte, J. W. M., et al., MRI Angiography of the Carotid Artery, Magn. Reson. Imaging,
Vol. 4, 1994, p. 497.
[159] Li, Z., L. Wei, M. Y. Gao, and H. Lei, One-Pot Reaction to Synthesize Biocompatible
Magnetite Nanoparticles, Adv. Mater., Vol. 17, 2005, p. 1001.
[160] Cinteza, L. O., et al., Diacyllipid Micelle-Based Nanocarrier for Magnetically Guided
Delivery of Drugs in Photodynamic Therapy, Molecular Pharmaceutics, Vol. 3, No. 4,
2006, pp. 415423.
98

[161] Ekimov, I. A., and A. Onushchenko, Quantum Size Effect in the Optical Spectra of Semiconductor Microcrystals, Sov. Phys. Semicond., Vol. 16, 1982, p. 775.
[162] Efros, L., and A. L. Efros, Interband Absorption of Light in a Semiconductor Sphere, Sov.
Phys. Semicond., Vol. 16, 1982, p. 772.
[163] Rossetti, R., S. Nakahara, and L. E. Brus, Quantum Size Effects in the Redox Potentials,
Resonance Raman Spectra, and Electrical Spectra of CdS Crystallites in Aqueous Solutions, J. Chem. Phys., Vol. 79, 1983, pp. 10861087.
[164] Brus, L. E., Electronic Wave Functions in Semiconductor Clusters Experiment and Theory, J. Phys. Chem., Vol. 90, 1986, pp. 25552560.
[165] Henglein, A., Small-Particle Research Physiochemical Properties of Extremely Small Colloidal Metal and Semiconductor Particles, Chem. Rev., Vol. 89, 1989, pp. 18611873.
[166] Alivisatos, A P., Semiconductor Clusters, Nanocrystals, and Quantum Dots, Science,
Vol. 271, 1996, p. 933937.
[167] Qu, L. H., and X. G. Peng, Control of Photoluminescence Properties of CdSe Nanocrystals
in Growth, J. Am. Chem. Soc., Vol. 124, 2002, p. 2049.
[168] Zhong, X. H., et al., Alloyed ZnxCd1xS Nanocrystals with Highly Narrow Luminescence
Spectral Width, J. Am. Chem. Soc., Vol. 125, 2003, p. 13559.
[169] Bailey, R. E., and S. M. Nie, Alloyed Semiconductor Quantum Dots: Tuning the Optical
Properties Without Changing the Particle Size, J. Am. Chem. Soc., Vol. 125, 2003, p.
7100.
[170] Bailey, R. E., A. M. Smith, and S. Nie, Quantum Dots in Biology and Medicine, Physica
E, Vol. 25, 2004, pp. 112.
[171] http://www.evidenttech.com.
Monodisperse CdE (E = S, Se, Te) Semiconductor Nanocrystals, J. Am. Chem. Soc., Vol.
115, 1993, p. 8706.
[173] Hines, M. A., and P. Guyot-Sionnest, Synthesis and Characterization of Strongly Luminescent ZnS-Capped CdSe Nanocrystals, J. Phys. Chem. B, Vol. 100, 1996, p. 468.
[174] Peng, X. G., et al., Epitaxial Growth of Highly Luminescent CdSe/CdS Core/Shell
Nanocrystals with Photostability and Electronic Accessibility, Am. Chem. Soc.,Vol. 119,
1997, p. 7019.
[175] Reiss, P., J. Bleuse, and A. Pron, Nano Lett., Vol. 2, 2002, p. 781.
[176] Taton, T. A., Nanostructures as Tailored Biological Probes, Trends Biotechnol., Vol. 20,
2002, pp. 277279.
[177] Salata, O. V., Applications of Nanoparticles in Biology and Medicine, J.
Nanotechnology, Vol. 2, 2004, p. 3.
[178] Bruchez, M., et al., Semiconductor Nanocrystals as Fluorescent Labels, Science, Vol. 281,
No. 5385, 1998, pp. 20132016.
[179] Chan, W. C., and S. Nie, Quantum Dot Bioconjugates for Ultrasensitive Nonisotropic
Detection, Science, Vol. 281, 1998, pp. 20162018.
[180] Mazzola, L., Commercializing Nanotechnology, Nature Biotechnol., Vol. 21, 2003, pp.
11371143.
[181] Paull, R., et al., Investing in Nanotechnology, Nature Biotechnol., Vol. 21, 2003, pp.
11341147.
[182] Sinani, V. A., et al., Collagen Coating Promotes Biocompatibility of Semiconductor
Nanoparticles in Stratified LBL Films, Nano Lett., Vol. 3, 2003, pp. 11771182.
[183] Goldman, E. R., et al., Multiplexed Toxin Analysis Using Four Colors of Quantum Dot
Fluororeagents, Anal. Chem., Vol. 76, 2004, p. 684.
[184] Han, M., et al., Quantum-Dot-Tagged Microbeads for Multiplexed Optical Coding of
Biomolecules, Nature Biotechnol., Vol. 19, 2001, pp. 631365.
99
[185] Tran, P. T., et al., Use of Luminescent CdSe-ZnS Nanocrystal Bioconjugates in Quantum
Dot-Based Nanosensics, Phys. Stat. Sol. B, Vol. 229, 2002, p. 427.
[186] Willard, D. M., and A. Van Orden, Quantum Dots: Resonant Energy-Transfer Sensor,
Nat. Mater., Vol. 2, 2003, p. 575.
[187] Clapp, A. R., et al., Fluorescence Resonance Energy Transfer Between Quantum Dot
Donors and Dye-Labeled Protein Acceptors, J. Am. Chem. Soc.,Vol. 126, 2004, p. 301.
[188] Wargnier, R., et al., Energy Transfer in Aqueous Solutions of Oppositely Charged
CdSe/ZnS Core/Shell Quantum Dots and in Quantum DotNanogold Assemblies Nano
Lett.,Vol. 4, 2004, p. 451.
[189] Bocsi, J., et al., Automated Four-Color Analysis of Leukocytes by Scanning Fluorescence
Microscopy Using Quantum Dots, Cytometry A., Vol. 69, No. 3, March 2006, pp.
131134.
[190] Le Gac, S. I. Vermes, and A. van den Berg, Quantum Dots Based Probes Conjugated to
Annexin V for Photostable Apoptosis Detection and Imaging, Nano Lett., Vol. 6, No. 9,
September 2006, pp. 18631869.
[191] Dubertret, B., et al., In Vivo Imaging of Quantum Dots Encapsulated in Phospholipid
Micelles, Science, Vol. 298, 2002, p. 1759.
[192] Gao, X. Y. et al., In Vivo Cancer Targeting and Imaging with Semiconductor Dots,
Nature Biotechnol., Vol. 22, No. 968, 2004.
[193] So, M. K., et al., Self-Illuminating Quantum Dot Conjugates for In Vivo Imaging, Nature
Biotechnol., Vol. 24, No. 3, March 2006, pp. 339343.
[194] Zimmer, J. P., et al., Size Series of Small Indium Arsenide-Zinc Selenide Core-Shell
Nanocrystals and Their Application to In Vivo Imaging, J. Am. Chem. Soc., Vol. 128, No.
8, March 1, 2006.
[195] Bakalova, R., et al., Quantum Dot Based Western Blot Technology was Used for
Ultrasensitive Detection of Tracer Proteins, J. Am. Chem. Soc., Vol. 127, No. 26, July 6,
2005, pp. 93289329.
[196] Kahn, E., et al., Analysis of CD36 Expression on Human Monocytic Cells and
Atherosclerotic Tissue Sections with Quantum Dots: Investigation by Flow Cytometry and
Spectral Imaging Microscopy, Anal. Quant. Cytol. Histol., Vol. 28, No. 1, February
2006, pp. 1426.
[197] Swift, J. L., R. Heuff, and D. T. Cramb, A Two-Photon Excitation Fluorescence CrossCorrelation Assay for a Model Ligand-Receptor Binding System Using Quantum Dots,
Biophys. J., Vol. 90, No. 4, February 15, 2006, pp. 13961410.
[198] Dahan, M., et al., Time-Gated Biological Imaging by Use of Colloidal Quantum Dots,
Opt. Lett., Vol. 26, 2001, p. 825.
[199] Kirchner, C., et al., Cytotoxicity of Colloidal CdSe and CdSe/ZnS Nanoparticles, Nano
Lett., Vol. 5, No. 2, 2005, pp. 331338.
[200] Derfus, A. M., et al., Probing the Cytotoxicity of Semiconductor Quantum Dots, Nano
Lett., Vol. 4, No. 1, 2004, pp. 1118.
[201] Hardman, R., A Toxicologic Review of Quantum Dots: Toxicity Depends on
Physicochemical and Environmental Factors, Environ. Health Perspect., Vol. 114 , No. 2,
February 2006, pp. 165172.
[202] Michalet X., et al., Quantum Dots for Live Cells, In Vivo Imaging, and Diagnostics, Science, Vol. 307, No. 5709, Jan. 28, 2005 pp. 53844.
[203] Gao, X., and S. Nie, Molecular Profiling of Single Cells and Tissue Specimens with Quantum Dots, Trends in Biotechnology, Vol. 21, No. 9, 2003, pp. 371373.
[204] Chan, W. C., Bionanotechnology Progress and Advances, Biol. Blood Marrow Transplant., Vol. 12, No. 2, Suppl. 1, January 2006, pp. 8791.
[205] Alivisatos, A.P., W. Gu, and C. Larabell, Quantum Dots as Cellular Probes, Annu. Rev.
Biomed. Eng., Vol. 7, 2005, pp. 5576.
100

[206] Lin, H., and R. H. Datar, Medical Applications of Nanotechnology, Natl. Med. J. India,
Vol. 19, No. 1, JanuaryFebruary 2006, pp. 2732.
[207] Gao, X., L.W. Chung, and S. Nie, Quantum Dots for In Vivo Molecular and Cellular
Imaging, Methods Mol. Biol., 2007.
[208] Caruthers, S. D., S. A. Wickline, and G. M. Lanza, Nanotechnological Applications in
Medicine, Curr. Opin. Biotechnol., Vol. 18, No. 1, February 2007, pp. 2630.

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CHAPTER 3

Nanoparticles for Biomedical Applications

Nanoparticles for Biomedical Applications

usually a few nanometers). As noble metals are reduced in size to tens of

3.2 Synthesis of Metallic Nanoparticles

or magnetic remanence synchronously. In other words, from the viewpoint of the

Synthesis of Metallic Nanoparticles

Synthesis Approaches to Noble Metal Nanoparticles

In fact, as mentioned earlier, since the seventeenth century, noble metallic

Nanoparticles for Biomedical Applications

In general, the process of generation of a solid phase from solution inevitably

Synthesis of Gold Nanoparticles

3.2 Synthesis of Metallic Nanoparticles

+ 3nB(OH) 4 + (32n + 8m)H + + 32nCl + 3nNa +

Nanoparticles for Biomedical Applications

3.2 Synthesis of Metallic Nanoparticles

implicating interactions between Au surface atoms and the nonbonding S orbitals of

Synthesis of Magnetic Metal Nanoparticles

Nanoparticles for Biomedical Applications

catalytic properties. In particular, the exponential dependence of the magnetization

3.2 Synthesis of Metallic Nanoparticles

superstructures possess lots of advantages in application in bionanotechnology and

Nanoparticles for Biomedical Applications

(CO) 8 ] 2Co + 18CO

3.2 Synthesis of Metallic Nanoparticles

Nanoparticles for Biomedical Applications

3.2 Synthesis of Metallic Nanoparticles

Nanoparticles for Biomedical Applications

3.3 Novel Properties of Metal Nanoparticles

Briefly, a three-neck flask with dichlorobenzene is heated to ~220C under N2

Novel Properties of Metal Nanoparticles

Unique Properties of Noble Metal Nanoparticles

Nanoparticles for Biomedical Applications

Three-dimensional superlattice structure of 7-nm cobalt nanoparticles. (Source: [79].)

3.3 Novel Properties of Metal Nanoparticles

In Whettens groups investigation, they measured the optical absorption spectra

Nanoparticles for Biomedical Applications

3.3 Novel Properties of Metal Nanoparticles

also to apply to Au clusters by cluster-beam experiments [94]. Now we discuss them

1.5 2.0 2.5 3.0 3.5 4.0

Nanoparticles for Biomedical Applications

1.5 2.0 2.5 3.0

3.3 Novel Properties of Metal Nanoparticles

Nanoparticles for Biomedical Applications

Here, p is the frequency of the plasma oscillation of free electrons expressed in

3.3 Novel Properties of Metal Nanoparticles

Nanoparticles for Biomedical Applications

3.3 Novel Properties of Metal Nanoparticles

employed, taking as adjustable parameters the number of electrons donated (or

Magnetic Properties of Metallic Nanoparticles

Nanoparticles for Biomedical Applications

Superparamagnetism. At room temperature, the nanoparticles usually exhibit

where K is the anisotropy constant, kB is Boltzmanns constant, and V is the average

3.3 Novel Properties of Metal Nanoparticles

Nanoparticles for Biomedical Applications

3.4 Application of Metal Nanoparticles in Biomedicine

not completely quenched is best explained by a partial coverage of the surface, in

Application of Metal Nanoparticles in Biomedicine

Biomedical Detection Using Novel Metal Nanoparticles

Functional nanomaterials that are designed to perform a reaction are of intense

Nanoparticles for Biomedical Applications

In the case of synthesis of 1-Pyren-1-yl-propyn-1-one (Pyr), an amount of

3.4 Application of Metal Nanoparticles in Biomedicine

Nanoparticles for Biomedical Applications