Mounting and labeling Flashcards
last step in tissue processing that results in a
permanent histological preparation suitable for
microscopy, after adhesion of the sections on to
the slide and appropriate staining of the tissue.
Mounting
usually a syrupy fluid applied between the section and the
coverslip after staining, setting the section firmly,
preventing the movement of the coverslip
Mounting medium
important because it governs the contrast between the
cellular detail and the background, and also the
transparency of the observed sample against the bright
field of the microscope
Refractive index
Mounting media may be divided
into two main groups:
- Aqueous Media
- Resinous Media
used for mounting
sections from distilled water when the stains would be
decolorized or removed by alcohol and xylene
Aqueous mounting medium
Aqueous mounting medium is made up of
made up of gelatin, glycerin jelly or gum arabic, glycerol, sugar, and a preservative solution.
Purpose of glycerin jelly or gum arabic
to
solidify the medium
Purpose of glycerol
to prevent cracking and
drying of the preparation
Purpose of sugar
to increase the
refractive-index
has a low refractive index, is moderately transparent and
evaporates easily, hence is good only for temporary
mounting.
Water
improves the image
quality and also supports the specimen.
Refractive index of water
Why wet mounts cannot be stored over extended time period
water will evaporate
has a high index of refraction and provides greater
visibility if slightly diluted with water
Glycerin
Glycerin refractive index
1.46
miscible with water, is inexpensive, and is non-poisonous.
Glycerin
Disadvantage of glycerin
- difficult to prepare slides that are truly permanent in
nature. - slowly run off a slide that is tilted.
- Do not stack slides for long as the pressure will squeeze
glycerin from the mounts. - Glycerin will eventually evaporate and air will penetrate
under the coverslip. - Glycerin can be attacked by microorganisms,
very suitable semi-permanent mounting medium
Glycerin
Commonly used to mount sections for immunofluorescence and glycerol may be added to other
agents to retard drying and cracking.
Phosphate buffered glycerol
Refractive index of Phosphate buffered glycerol
1.47
Formula of GLYCERIN JELLY (KAISER’S
1880)
- Gelatin 10 gm.
- Glycerol 70 ml.
- Distilled water 60 ml.
- Phenol crystals (preservative) 0.25 gm.
standard mounting medium used
when dehydration and clearing with xylene cannot be
made (as in fat stains)
Glycerin jelly
Glycerin jelly refractive index
1.47
FARRANT’S MEDIUM formula
- Gum arabic 50 gm.
- Distilled water 50 ml.
- Glycerol 50 ml.
- Sodium merthiolate 0.025 gm
FARRANT’S MEDIUM. Refractive index
1.43
Advantage of farrant’s medium
does not solidify upon storage and therefore does not
need to be heated before use.
Disadvantage of farrant’s medium
takes a longer time to harden and may therefore require
ringing
Arsenic trioxide may be used as a substitute of sodium
merthiolate for preservation of the medium.
Farrant’s medium
APATHY’S MEDIUM formula
- Pure gum arabic (crystals not powder) 50 gm.
- Pure cane sugar or sucrose 50 gm.
- Distilled water 50 ml.
- Thymol crystals 0.05 gm.
Apathy’s medium refractive index
1.52
used for methylene blue-stained nerve preparations and
as a general purpose aqueous mountant.
Apathy’s medium
Advantage of apathy’s medium
one of the most useful aqueous mountants for fluorescent
microscopy, being virtually non- fluorescent.
Disadvantage of apathy’s medium
sets quite hard, has a higher refractive index, and does not
require ringing.
recommended for mounting frozen
sections from water.
BRUN’S FLUID
BRUN’S FLUID formula
- Glucose 24 gm.
- Glycerine 6 ml.
- Spirits of camphor 6 ml.
- Distilled water 84 ml.
used for preparations that have been
dehydrated and cleared in xylene or toluene, and are
recommended for majority of staining methods.
RESINOUS MOUNTING MEDIA
RESINOUS MOUNTING MEDIA is divided into 2
natural and synthetic resins.
most important synthetic resins are used for
embedding undecalcified bones,
and for electron
microscopy
It is a transparent, almost colorless oleoresin that adheres
firmly to glass and sets to a hard consistency without
granulation.
Canada Balsam
Canada Balsam refractive index
1.524
recommended for whole mounts and
for thick sections because it does not shrink much.
Canada balsam
may be added to Canada balsam to maintain its
neutral reaction
Calcium carbonate chips
dissolved in xylene in an incubator at 37°C or paraffin
oven at 58 °C
Canada balsam
Canadian tree:
Abus Balsamea
recommended for small tissue sections but not for whole
mounts because of shrinkage produced on drying
DPX - (Dibutyl Phthalate and
Xylene)
Disadvantage of DPX - (Dibutyl Phthalate and
Xylene)
It tends to set quickly and often retract from
the edge of the coverslip.
colorless, neutral medium in which most standard stains
are well preserved.
DPX - (Dibutyl Phthalate and
Xylene)
DPX - (Dibutyl Phthalate and
Xylene) refractive index
1.532
synthetic resin mixture in xylene, available in a
pale yellow or colorless solution
XAM
Xam refractive index
1.52
Dries quickly without retraction, and preserves stains
well.
Xam
CLARITE refractive index
1.544
synthetic resin which is soluble in xylene
generally preferred over D.P.X
CLARITE
Other recommended
synthetic mounting
media
- Permount (made by Fisher Scientific)
- H.S.R. (Harleco Synthetic Resin)
- Clearmount (Gurr)
process of sealing the margins of the cover-slip to prevent
the escape of fluid or semi-fluid mounts and evaporation
of mountant, to fix the coverslip in place, and to prevent
sticking of the slides upon storage.
Ringing
The term “ringing” originated because
round coverslips
were initially used and the coating applied in the form of
a circle or “ring.”
made up of two parts paraffin wax
mixed with 4-9 parts powdered colophonium resin,
heated and filtered.
Kronig cement
cellulose adhesives
Durofix
often used as a mountant in
immunofluorescence microscopy, has been
recommended as an alternative for glycerin jelly
Polyvinyl alcohol
