Mounting Media

Dr Iram Iqbal
Aim
To define and discuss different type of
mounting medias ,their technique for
mounting sections ,and the qualities
of the ideal mountant.
 sequence
 Defination
 Qualities of an ideal mountant
 Classification of mounting media
 Cover glasses
 Properties of the good mounted sections
 Technique for mounting sections
 Labelling & and storage of slides
MOUNTING OF
SECTIONS
 After staining and before
examination, sections should be
mounted with a suitable medium under
cover slips .
 To facilitate ease of handling and
storage and to prevent damage to
the section.
 The liquid used for mounting slides is
known as “Mountant”.
 Qualities of an ideal
mountant
 An ideal mounting medium must fulfill
following criteria;
 It should fulfill the basic

requirement of protecting from

dust and damage.
 It should preserve the cell
characteristics and quality of the
stain.
 It should be colourless and

transparent .
AN IDEAL MOUNTANT
 The medium should have
a refractive index as
close as possible to that
of the glassi,e,; 1.5 .
 It should not cause
stains to “bleed”or fade
 It should not crack or
appear granular on
setting.
 It should be non sticky
and harden quickly.
Cont…
 It should not shrink back from the
edges of the cover glass.
 It must be free flowing.
 It must be free from bubbles.
 It should be resistant to
contamination
 Once set, the mountant should remain
stable.
Types of mounting Media

 Aqueous based mounting media

 Resinous mounting media .

AQUEOUS MOUNTANT

 Apathy,s Medium (RI 1.52)

 Farrant,s Medium (RI 1.43)

 Fructose (Levulose) Syrup (RI 1.47)

 Glycerin Jelly (RI i.47)

 Highman,s Medium (RI 1.52)

RESINOUS MOUNTANTS
 Canada balsam (RI 1.52)
 Cover bond (RI I.53)
 DePeX (RI 1.52)
 Gurr,s Neutral Mounting Medium(RI 1.51 )
 Histoclad( RI 1.54)
 Permount (RI 1.526)
 Pro-Texx (RI 1.495)
 Technicon Resin (RI 1.62)
 UVinert (RI 1.517)
 XAM (RI 1.52)
It is important to note also that
the clearing agent used must be
compatible with the mounting
medium, or the sections must be
thoroughly dried prior to
mounting.
 Aqueous Mounting Media
Aqueous mounting media are used for
mounting sections from distilled
water.
 AQUEOUS MOUNTING
MEDIA
 It is used in cases where the stain
would be decolourized or removed by
alcohol and xylene
 Aqueous mountants require the addition
of a bacteriostatic agent such as
phenol to prevent the growth of molds
 Once the medium has hardened, a
permanent seal can be obtained by
ringing the coverglass with clear nail
polish
Apathy’s Medium (RI 1.52)
It is a very useful medium for mounting
sections for
fluorescent microscopy.
 Gum Arabic 50g
 Sucrose 50g
 Distilled water 50ml
 Thymol 1 crystal
This medium remains fluid in storage and
hardens by evaporation .
Should be stored in well stoppered bottle.
 Farrant`s Medium (RI1.43)
Gum Arabic 50g
Glycerin 50ml
Distilled water 50ml
Sodium merthiolate 0.025g

This medium remains fluid in storage

there for more convenient than
glycerin jelly.
Its disadvantage lies in the fact that It
takes longer to set. Store in a well
stoppered bottle.
Fructose (Levulose) Syrup (RI 1.47)
Recommended as a temporary mountant and for
special techniques.
 Fructose 75g
 Distilled water 25ml
Glycerin Jelly (RI 1.47)
An excellent routine mountant for fat stains.
 Gellitin 10g
 Distilled water 60ml
 Glycerin 70ml
 Phenol 0.25g
The gelatin causes this medium to solidify and it
must be melted before use.
 This is easily done by placing it in the oven
or a water bath at 60 C Sometime prior to
use.
 Do not shake or stir the medium during
melting as this will produce bubbles that are
difficult to remove.
Highman`s Medium (RI 1.52)
This medium is recommended for use with
the metachromatic dyes especially methyl
violet.
 Gum Arabic 20g
 Sucrose 20g
 Potassium acetate 20g
 Sodium merthiolate 10ml
 Distilled water 40ml
This medium remains fluid in storage and
hardens by evaporation. Store in a well
stoppered bottle.
 RESINOUS MOUNTING
MEDIA
 Resinous mountants are either
natural or synthetic resins
dissolved in solvents such as
benzene, toluene or xylene.
Canada balsam
 It is very expensive
natural resin.this is
prepared by collecting
the resin exuded by
Abies Balsamica (the
Balsam fir) and dilute it
in the solvent eg,
xylene.
Canada balsam
 As canada balsam does not mix with
water mounting in it implies the use of
sequence of dehydration,starting with
 Low grade alcohols followed by
 High grade alcohol
 Absolute alcohol
 Mixed cleaning agent plus alcohol
 Cleaning agent
 Cleaning agent mixed with xylene
 Pure xylene and
 Balsam dissolved in xylene.
 toluene or benzene could be used instead
of xylene ,but these solvent are equally
toxic and dangerous.
 Development of some synthetic is
substitutes for balsam doesn't solve the
problem. they are equally expensive, that
need the same steps and use the same
toxic solvents.
 There are less toxic and less dangerous
substances but they are expensive.
 Canada balsam has
an RI of1.52 very
near of the glass
slides and cover slips
 The mounting media
must always have an
RI higher than the
mounted sample.
Canada balsam (RI 1.52)
This may be purchased as a readymade
solution of 60% resin by weight in xylene or
in the dry form.
 If it is obtained in the dry form, it is made

up as a 55 – 70% solution in xylene.

 A few chips of marble in the bottom of the

bottle will help reduce acidity.

Coverbond (RI 1.53)
This is Harleco Synthetic resin (H.S.R.)
dissolved as a 60% solution by weight in
toluene.it may be purchased s dry resin and
may be dissolved in toluene or xylene.
DePeX (RI 1.52)
Gurr`s DePeX is a polysterene resin dissolved as a
20% solution in xylene. Because this mountant
tends to shrink on hardening, it should be added to
excess. Overflow is easily removed with a scalpel
after drying is complete.
Gurr`s Neutral Mounting Medium (RI 1.51)
This is a mixture of coumarone and other resins as
76% solution in cineol. This mountant is a rather
viscous solution.
Histoclad (RI 1.54)
Clay Adam’s Histoclad is a 60% solution of a
synthetic resin in toluene.
Permount (RI 1.528)
Fisher’s Permount is a 60% solution of naphthalene
polymer in toluene.
Pro-Texx (RI 1.495)
Pro-Texx from Lerner Laboratories is a
mounting medium of neutral pH and with an
antioxidant additive to preserve stain quality.
It is soluble in both toluene and xylene.
Technicon Resin (RI 1.62)
This mountant from the Technicon
Corporation is a 60% solution of couramone-
indene polymer in benzene and xylene in equal
parts.
It has a tendency to form bubbles on
hardening due to high volatility of benzene.
UVinert (RI 1.517)
A patented mounting medium that is non
fluorescent from Gurr.
Properties Of Good Mounted
Sections
 Present a neat appearance.
 There should be no excess mountant
on or around the coverglass.
 There should be no bubbles under
the cover glass.
Usual causes of messy slides
are
 Too much mountant
 Too thick a mountant that has not
been spread to the edges of the
cover glasse
 If the mountant is too thick it can be
diluted with either toluene or xylene.
 Coverglasses
 Glass coverslips are used to protect
specimens from physical damage and to
keep them from drying out when exposed to
air after staining.
COVERGLASSES
 Coverglasses are
either square or
rectangular
 Glass coverslips are
used to protect
specimens from
physical damage
 They come in
different sizes and
widths
Cont…
 They are either
22mm or 24mm
wide and come in
four different
lengths such as,
22mm, 30mm,
40mm or 50mm
 Coverglasses
 Coverglasses are manufactured to a
specified thickness to complement the
optical specifications of microscope
objectives lenses(the recommended
thickness is indicated on the barrel of the
lens and is normally 0.17 mm).
 Various thicknesses are available (with some
variation between manufacturers) and are
designated
Cont…
 Various thicknesses are available
 No. 1 (0.13 - 0.17 mm)
 No. 1½ (0.16 - 0.19 mm)
 No. 2 (0.19 - 0.25 mm)
 TECHNIQUE FOR
MOUNTING SECTIONS
 Select the appropriately sized coverslip and
place on a white paper sheet.
 Place a drop of mountant in the centre of the
coverslip
 Invert the slide (section face down) over the
coverslip and with one end resting on the
paper sheet; gradually lower the other end
until the mountant touches the section.
 Mountant will spread quickly over the section,
between slide and coverslip.
Cont…
 The slide, with coverslip attached, is then
turned upright.
 Any trapped air is gently squeezed out
whilst aligning the coverslip.
 The mountant is allowed to set.
 The time required will depend upon the
particular agent used.
 If the result is inadequate, slides are
returned to the solvent (appropriate to the
mountant) to have the coverslip removed
and the process repeated
Other techniques of
mounting
 Place the slide on a
level surface, and apply
a drop of mountant
using the dispenser rod
 Hold the cover slip at a
45° angle to the surface
of the slide, and allow
the bottom edge to
touch the drop of
mountant. When the
drop has spread along
the edge of the slip, let
go of the slip and allow
the Histomount to
spread slowly
Cont….
 The stained section
on the slide is
mounted under a
thin glass coverslip.
The technician
takes care to keep
any bubbles from
being trapped in
the resinous
mounting medium.
Cont…
 Excess mounting medium may be
removed while wet with a tissue, or
with a razor blade when dry.
 Histomount will dry sufficiently to
be read in 30 minutes.
LABELLING & STORAGE OF
SLIDES
 After staining and
mounting apply a
paper label with:
 Name of tissue
 date
 laboratory number
 staining method
REFERENCES:
 Theory and Practice of Histological Techniques
John D Bancroft & Marilyn Gamble
Fifth Edition
5. An Introduction to Histotechnology
Geoffrey G. Brown, AIMLS