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MINIATURISATION FOR CHEMISTRY, BIOLOGY & BIOENGINEERING
Electroosmotic mixing in microchannels{
Ian Glasgow, John Batton and Nadine Aubry
Department of Mechanical Engineering, New Jersey Institute of Technology, Newark,
NJ, USA
Received 15th June 2004, Accepted 24th September 2004
First published as an Advance Article on the web 4th November 2004
Published on 04 November 2004. Downloaded by University of Texas Libraries on 19/10/2014 07:00:38.
Mixing is an essential, yet challenging, process step for many Lab on a Chip (LOC) applications. This paper
presents a method of mixing for microfluidic devices that relies upon electroosmotic flow. In physical tests and
in computer simulations, we periodically vary the electric field with time to mix two aqueous solutions. Good
mixing is shown to occur when the electroosmotic flow at the two inlets pulse out of phase, the Strouhal
number is on the order of 1, and the pulse volumes are on the order of the intersection volume.
Introduction
Mixing is a crucial process in many microfluidic systems such
as micro Total Analysis Systems (microTAS), Lab on a Chip,
and microreactors which require that reagents be added and
samples diluted. Unfortunately, at this size scale and with
liquids flowing relatively slowly, the flow is laminar and
confluencing liquids tend to flow side by side.
Because of the importance of mixing in microfluidics
systems, numerous researchers have developed various mixing
techniques, based upon geometries that induce secondary
flow1,2 or reduce diffusion length scales,3 incorporate miniature
mixing rods or balls,4 utilize cross flows5 or alternating flow
from the inlets,6 pulse one of the reagents,7 or apply magnetic,8
electric,9,10 or ultrasonic vibratory11 fields. Our earlier work12
demonstrated that pulsed flow, generated by mechanically
displacing the liquid, such as by periodically reversing the
pumps, could cause two aqueous solutions to mix. While this is
a simple method of mixing, many systems rely upon
electroosmotic pumping, which uses electric fields instead of
mechanical pumps.
Electroosmotic flow is an elegant method for flow control in
microfluidic and nanofluidic systems since an electric field,
instead of moving parts, controls the flow. Electrodes are
generally easier to fabricate and incorporate in these miniature
devices than are pumps and valves. In this paper we show that
mixing can be achieved completely electroosmotically. Oddy
et al. in ref. 10 studied electrokinetic mixing with two setups:
one in which the electric field applied to the main channel
varied with time, thus varying the flow rate of both fluids in
unison, after the confluence; and one in which the time varying
electric field is applied to a cross-channel perpendicular to the
main channel. In contrast, our technique which uses
the application of pulsed voltages to the electrodes located in
the reagent wells, causes the two fluids to differentially vary
their flow rates from the inlet channel to the confluence. We
demonstrate our method via computer simulations and
physical models.
Experimental setup
Physical tests
DOI: 10.1039/b408875a
Computer simulations enable easy control of the flow
parameters and extensive data collection at all locations in
the channel while physical microchannel devices enable
{ Electronic supplementary information (ESI) available: Electroosmo-
tic flow mixing. See http://www.rsc.org/suppdata/lc/b4/b408875a/
558 Lab Chip, 2004, 4, 558562 This journal is The Royal Society of Chemistry 2004
verification testing. The physical device consists of a 6.5 mm
thick polycarbonate base, a 0.5 mm thick polycarbonate cover,
and three 6.5 mm tall polycarbonate wells. Fig. 1 shows the
channel geometry. The base contains two 120 mm deep by
200 mm wide by 6 or 8 mm long troughs, which form the T
shaped channel network. The troughs were machined with a
0.006 inch diameter carbide end mill (Ultra-Tool, Huntington
Beach, CA). The cover contains three access holes for the wells,
each 3 mm in diameter. A UV curing acrylic adhesive (NOA 72,
Norland Products, Cranbury, NJ) is placed on the base before
the cover is set on top. The adhesive is allowed to wick between
the base and cover for complete coverage. To help prevent
the adhesive from wicking into the troughs, the edges of the
troughs are kept sharp. Curing the adhesive with a UV lamp
firmly bonds the cover to the base. The three wells, which each
consist of a small block of polycarbonate with a 6 mm hole
through the thickness, are bonded to the cover with epoxy.
In order to setup the electric field in the channel, 1 mm
diameter platinum electrodes are placed in each of the wells.
The electric field takes the form of a biased squarewave (as
shown in Fig. 1) or biased sinusoidal wave. The bias or
constant DC voltage between both inlets and the outlet,
generates the base or net flow. The periodic component that
varies at frequencies of 0.5 Hz to 10 Hz, between each inlet and
the outlet, causes the pulsed flow. In these tests, either 42 or 60
VDC bias, from a series of batteries, is applied between the
inlets and outlet to generate the base flow. Two types of time
varying voltage components between the inlets and the outlet
are tested: squarewaves at 180u out of phase or sinusoids at 90u
out of phase. The earlier tests use squarewaves of several
hundred volts peak to peak. Two high voltage amplifiers (Trek
Inc., NY) have a small constant voltage as an input and their
outputs are sent through a switched double pole double throw
relay to the electrodes in the inlet wells. A signal from the
function generator controls the relay, periodically reversing
which amplifier drives which electrode. The sinusoidal wave-
form voltages are generated in a different manner. A function
generator sends out a sinusoidal control signal to the first
amplifier and in parallel to a small phase delay circuit. The
output from the phase delay circuit goes to the second
amplifier. The outputs from the amplifiers go directly to the
two inlet port electrodes. The fluid levels in the three wells are
allowed to equilibrate before the voltages are applied. In
comparison to many microfluidic devices, these channels are
relatively short and have large cross-sections, so they exhibit
less resistance to flow, are more sensitive to liquid level
differences, and require more time to equilibrate.
Before each use, the channels are cleaned with 0.18 M NaOH
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Fig. 1 Geometry of the device showing the voltages applied to the electrodes for the case of square wave voltages with an 180u phase difference at
the two inlets.
and rinsed with deionized water. Both inlet wells are filled with
similar aqueous solutions. One well contains deionized water,
the other deionized water with a minimal amount of rhodamine
dye for visualization purposes. In some tests, acetate buffer, at
a concentration of 40 mM, held the pH at a value of 8.3. A
fluorescence microscope (Nikon MEC 600 Eclipse with EPI-
Fluorescence attachment) enables observation of lateral
mixing. Three-dimensional mixing at all locations in the
channel is studied in more detail using computational fluid
dynamics.
Several basic measurements were taken. Measuring the
current at a given voltage, and conservatively assuming that no
heat escapes through the cartridge, indicates the power
dissipated in the fluid. Measuring the velocity of the fluid at
a voltage enables the calculation of the electroosmotic mobility.
The base flow velocity, V B, through the outlet channel ranges
from about 0.05 to 0.3 mm s21. The outlet channel has a length
scale, L, defined by the hydraulic diameter as 4 times the area
divided by the perimeter, or 0.15 mm in this case. The
kinematic viscosity, n, of water at room temperature is
approximately 1026 m2 s21. Thus, the Reynolds number
(Re ~ V BL/n) is about 0.08, which indicates that the flow is
very laminar, i.e., 4 and a half orders of magnitude below the
transition to turbulent flow. The Schmidt number, Sc, is the
ratio of the kinematic viscosity to the mass diffusivity, D (Sc ~
n/D). For a typical application such as mixing an aqueous
solution containing small proteins, D ~ 10210 m2 s21 and Sc
takes the value of 104. This latter ratio is very high due to the
fact that the kinematic viscosity for such solutions is much
larger than the mass diffusivity. The Peclet number, Pe, the
product of the Reynolds and Schmidt numbers, is often used as
a measure of diffusive mixing. Pe ~ Re 6 Sc ~ VL/D. In this
case, the Peclet number is about 0.8 6 103, which indicates that
the time required for diffusive mixing is quite large.
While the simulations provide concentration data through-
out the channels at all times, obtaining such data from physical
experiments while the pulsing occurs is not practical. In
particular, we fall short of providing true three-dimensional
data on a linear scale based on dye intensity measurement.
Numerical simulations
Computational fluid dynamics simulations are invaluable
because they enable data collection at all points in the model
at all times during the pulse cycles. The simulations are
conducted using Fluent (Fluent Inc., Lebanon, NH). The
simulations model the region of the device near the confluence:
1.25 mm long sections of inlet channels and a 3 mm section of
the outlet channel. The mesh is discretized into hexahedral cells
with 10 mm sides (width, depth, and height). Near the ports, far
from the intersection and mixing region, the cells are 10 by 10
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by 50 mm long along the flow direction. The model begins with
steady flow at the base flow rate, then proceeds with 20 time
steps per cycle. Previous tests12 verified the accuracy of the
model. In electroosmotic flow, the fluid is propelled in the
Debye layer along the channel walls. Across this layer the fluid
velocity varies from no-slip at the wall to the maximum
electrically induced velocity on the other side of the layer.
However, because this layer is typically on the order of
nanometers, i.e., 4 to 5 orders of magnitude less than the
characteristic size of the channels, the thickness of this layer is
negligible. Accounting for the velocity gradient in this layer
would not significantly change the results but would substan-
tially increase the computational complexity, so the velocity
gradient in this extremely thin layer is disregarded, as in refs. 13
and 14.
The electroosmotic flow is thus modeled by specifying the
fluid velocity at the walls, in a manner similar to that described
by Ermakov et al.13 and by Qiao and Aluru.14 As outlined in
ref. 13 the velocity along the walls, V, may be modeled
according to the following equation
V ~ mEOE (1)
where mEO is the electroosmotic mobility and E is the electric
field intensity in the longitudinal direction. This expresses that
in an electroosmotic flow, the velocity of the liquid one Debye
length from the walls is proportional to the electric field
intensity. To determine the fluid velocity at the walls, which
varies spatially, particularly near the confluence, the electric
fields are modeled by solving the Laplace equation for the
electric potential using ANSYS (ANSYS, Inc., Canonsburg,
PA). The electric field values are multiplied by a constant and
then imported into Fluent such that the velocities at the walls
are set to be proportional to the electric field in the X and Y
directions. The electric field in the Z (depth) direction is found
to be orders of magnitude smaller and is thus neglected.
The flow simulations are first order models that do not
account for higher order effects such as viscosity changes due to
Joule heating or chemical reactions.
Both the inlet liquid streams are chosen as aqueous solutions,
modeled as water at room temperature, with a viscosity of
0.001 m2 s21 and a density of 1000 kg m23. Diffusion is an
important aspect of mixing that must be modeled. A diffusion
constant of 1 6 10210 m2 s21, typical of the diffusion of small
proteins in aqueous media and other microfluidic applications
is used.
To evaluate the extent of mixing, we statistically analyze the
concentration of the liquid from one of the inlets at all cells in a
cross-section 0.5 mm downstream of the confluence (see also
ref. 12). This location is a reasonable distance (two channel
widths) from the confluence zone and a reasonable place for the
Lab Chip, 2004, 4, 558562 559

next station in a micro total analysis system. For simplicity, the
base flow rate from both inlets is set to be the same so that the
ideal concentration, i.e. for a completely mixed solution,
should be 0.50 in every cell. The deviation about a mean would
yield the variation in concentration; a value of 0 would indicate
perfect mixing. We modify the deviation about a mean to
weight each cell by its flow rate and normalize the equation so
that the values range from 0 (complete absence of mixing) to 1
(complete mixing). The degree of mixing is given by,
s fields or mechanical pumping.
 
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Pn
(xi {
)2 qi
n qmean
i~1
degree of mixing~1{

Numerical simulations reveal the benefits of pulsed flow
where n represents the number of cells in the cross-section, xi is
the concentration in the ith cell, m is the mean concentration
which equals 0.50, qi represents the flow rate in the ith cell and
qmean is the mean flow rate of all the cells.
Results and discussion
Physical tests
Several basic measurements were taken to evaluate the
electroosmotic flow. Measurements of fluid velocity for five
values of the electric field indicate that the electroosmotic
mobility, as determined in eqn. (1) has the value of 6.0 6
1028 m2 V21 s21, with a coefficient of variation less than 11%.
When the electric field in the outlet channel is 187 V cm21,
1.4 mW are dissipated in the fluid and cartridge. Conservatively
assuming no heat is absorbed by the cartridge, then the fluid,
which is flowing at 27 nL s21, will heat up by a maximum of
12.4 uC. In practice, heat will transfer into the cartridge and
then into the surrounding air, so the fluid would not heat up as
much. Naturally, if the voltage were only applied for a short
time, perhaps 30 s, and assuming most of the heat would
transfer into the liquid in the cartridge, then the Joule heating
would cause less than a 5 uC rise in temperature.
Most of the physical tests were conducted with a base flow
mean velocity of 0.3 mm s21. The tests demonstrate that under
steady voltage conditions the two solutions flow side by side, as
shown in Fig. 2a and d. As expected, in this case there is
fluorescence in only half the outlet channel. In contrast, while
the electric field is pulsed at either 8 Hz or 10 Hz with the inlets
pulsing at 180u or 90u out of phase, the rhodamine solution fills
the outlet channel, as shown in Fig. 2b, c, e and f. This
demonstrates that the liquids are no longer restricted to their
respective sides of the channel. Fig. 2b and c are images taken
Fig. 2 Photographs of the electroosmotic flow in the T channel for various applied voltages at the inlets: (a) flow under steady voltage (initial
condition for b and c); (b) and (c) flow under time varying voltages at two instants during a 10 Hz square wave pulse cycle, with the inlets pulsing 180u
out of phase; (d) flow under steady voltage (initial condition for e and f); (e) and (f) flow under time varying voltages at two instants during a 8 Hz
sinusoidal cycle, with the two inlets pulsing 90u out of phase.
560 Lab Chip, 2004, 4, 558562
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half a pulse cycle apart, as are Fig. 2e and f. Notice the
differences in the confluence region. Tests were also conducted
with the smaller base flow mean velocity of 0.15 mm s21,
showing better mixing. This is consistent with the improvement
of mixing for higher Strouhal number values, as reported in ref.
15 for pressure driven flows.
The out of phase pulsing causes the liquids to mix,
independently of how the base flow is generated, i.e. the
mixing occurs whether the base flow is generated by electric
Numerical simulations
(2)
mixing. For instance, when the fluid is flowing through the
outlet channel at 2.0 mm s21, i.e. a flow rate of 60 nl s21, as
in the physical tests described above, superimposing 8 Hz
squarewave pulsing with a mean velocity of 4 mm s21 at both
inlets, with a 90u or a 180u phase difference between the two
inlets, leads to a substantial improvement in mixing. Without
the pulsing, mixing is predominantly due to diffusion, and the
degree of mixing is about 0.04. However, with the pulsing, the
degree of mixing is about 0.34 for both 90u and 180u phase
differences.
When the base flow is slower, sub-volumes of liquid spend
more time traveling through the confluence region and are thus
subjected to more pulses while in the confluence region, thus
leading to an increased degree of mixing. For instance, when
the base flow in the current case is decreased from 2.0 to
0.8 mm s21, the degree of mixing substantially increases from
0.34 to 0.80 and 0.59 for 90u and 180u out of phase pulsing,
respectively.
Fig. 3 represents the degree of mixing as a function of
accumulated volume, beginning with the transition from steady
flow without pulsing to the initiation of superimposed pulsing.
In the case of a phase difference of 180u out of phase pulsing,
while the electric potential applied at one inlet well is at a peak,
the potential at the other inlet is at a trough, which leads to no
net change in the flow rate through the outlet channel. The
degree of mixing versus accumulated volume curve is sigmoidal
shaped. The first part of this curve is nearly flat before the
accumulated fluid volume reaches about 15 nl. Indeed, there is
little evidence of mixing at a plane 0.5 mm downstream of the
center of the intersection until the mixed portion, which travels
at the base flow velocity, reaches this plane. At a mean velocity
of 0.8 mm s21, corresponding to a flow rate of 24 nl s21, 0.63 s
will elapse, corresponding to an accumulated flow of 15 nl after
the onset of pulsing. After this time, mixing improves as the

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Fig. 3 Degree of mixing as a function of accumulated fluid volume for
pulsing superimposed upon a base flow of 0.8 mm s21, showing that the
simulations predict superior mixing performance with 90u out of phase
pulsing.
fluid which has undergone more pulses in the confluence region
reaches this plane. Approximating the most consequential
region for mixing as extending about two channel widths
upstream of the intersection zone, the mixing will substantially
improve for the next 30 nl of volume (up to an accumulated
volume of 45 nl). Fluid upstream of this confluence region
when the pulsing initiates will undergo the largest number of
pulses as it travels through the confluence region. As this fluid
reaches our measurement plane, 0.5 mm downstream of the
confluence, the degree of mixing levels off.
In contrast, with 90u out of phase squarewave pulsing, the
net flow through the outlet channel is not steady, but oscillates
at the frequency of the pulsing. Portions of the fluid from either
of the inlet channels extend temporarily into the outlet channel,
and are then withdrawn, thus leading to some downstream
mixing not present in the 180u case. Thus, the corresponding
degree of mixing curve in Fig. 3 has practically no initial level
region; the extent of mixing begins to rise almost immediately.
The additional stretching and folding of the interface in this 90u
out of phase pulsing case leads to greater mixing. In the earlier
case of the greater base flow of 60 nl s21 (2.0 mm s21), the fluid
is not sufficiently withdrawn back through the confluence zone
to yield an improvement over the 180u phase difference case.
Fig. 4 Mass fraction contour levels at mid-depth of the T channel and in a cross-section 0.5 mm downstream of the center of the intersection for the
electroosmotic flow at a rate of 24 nl s21 through the outlet channel: (a) under steady voltage, (b) under time varying voltages at the inlets consisting
of a constant (base) and a superimposed pulsing at 8 Hz whose amplitude is ten times as large as that generating the base flow at the two inlets. There
is a 90u phase difference in the square waves at the two inlets in this simulation.
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Fig. 4 displays contour plots of the concentration of the fluid
from the lower inletat half the channel depth and at a cross-
section 0.5 mm downstream of the center of the intersection.
Comparing Fig. 4a, representing no pulsing, with Fig. 4b,
where an 8 Hz pulsing is imposed, with applied voltages at the
two inlet electrodes 90u out of phase, we notice the dramatic
increase in mixing due to the pulsed electric field. The
concentration is somewhat independent of the channel depth,
showing that pulsed electrokinetic mixing in this case is quasi
two-dimensional.
From tests and simulations entailing pressure driven and
displacement driven pulsing, the Strouhal number and the
pulse volume ratio are found to be the salient parameters.15
The Strouhal number, which is used to characterize fluidic
systems where a frequency is present, is the ratio of the
characteristic time period of the flow to the forcing (pulsing)
period.
L=VB fL
St~ ~ (3)
1=f VB
For example, when the pulsing frequency f is 8 Hz and the
base mean velocity V B ~ 2.0 mm s21 in the outlet channel
where L, the hydraulic diameter, is 150 mm, the Strouhal
number equals 0.6. The Strouhal number can be increased to
1.5 by decreasing the base flow to 0.8 mm s21 or by increasing
the pulsing frequency to 20 Hz. Higher Strouhal numbers
correspond to sub-volumes of fluid experiencing more pulses,
and therefore likely better mixing. However, if the pulse
frequency is too high, the fluid will not approach fully
developed flow during each pulse, which may adversely
affect the mixing. The Stokes number, which is the ratio of
the characteristic time period to establish fully developed flow
to the forcing period, is also the product of the Reynolds and
Strouhal numbers. For Stokes numbers significantly above a
value of one the flow will vary substantially from fully
developed flow. For example, at a base flow of 0.8 mm s21 and
a hydraulic diameter of 150 mm, a frequency of 44 Hz leads to a
Stokes number of 1.0, so the flow will vary substantially from
fully developed at frequencies several times greater.
We define the pulse volume ratio (PVR) to be the ratio of the
displaced volume during a pulse to the volume of
the intersection region in the T confluence, that is the
rectangular piped region defined by the width of the inlet
channels (0.2 mm), the width of the outlet channel (0.2 mm),
Lab Chip, 2004, 4, 558562 561

and the depth of the channels (0.12 mm). The pulse volume can
be approximated as the pulsed flow rate integrated over the
time period for half a pulse cycle. In the two electrokinetic flow
cases just described, the pulse volume is approximately 0.25 mm
of channel, while the volume of the intersection region is
equivalent to 0.2 mm of channel, yielding a ratio of 1.25.
pulse volume
PVR~ (4)
intersection region
When PVRs are significantly less than 1, each pulse forces
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only a very small subvolume of liquid back and forth through
the confluence region. However, if the PVR is too large, the
pulse volume will extend well into the inlet and outlet channels.
Optimal values for the PVRs are in the range of about 1 to 3.
Other methods of forcing the fluid to flow, such as electro-,
magneto-, and electromagneto-hydrodynamic pumping should
also enable pulsed flow mixing, but were not studied in this
paper.
Conclusion
Mixing, which is needed in many microfluidics systems, can be
readily accomplished simply by pulsing the flow from both
inlets, out of phase with each other, in electroosmotically
driven flow systems. Beyond the right selection of pulsing
parameters and a method to regulate the pulsing, nothing else is
required to implement the mixing technique. Any EOF driven
system can incorporate mixing, just by making the voltage vary
with time.
Acknowledgements
The authors are grateful to the New Jersey Commission on
Science and Technology for their financial support under
562 Lab Chip, 2004, 4, 558562
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Grant No. 01-2042-007-25, through the New Jersey Centre for
Micro-Flow Control and to the W. M. Keck Foundation for
their grant to establish the NJIT W. M. Keck laboratory. We
are indebted to John Hoilowski for his machining skills and to
Gianni Borghesan, Lee Carlson and George Bernes for
providing laboratory assistance.
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