CHAPTER 15 ADHESIVES AND MOUNTING MEDIA Flashcards
is the 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
After draining, the sections are fixed to the slides. This can be done either by leaving the slides in a [?] incubator overnight, by placing the slides in a wax oven at [?], or by drying the slides on a hot plate at [?].
37°C
56° to 60°C for 2 hours
45° to 55°C for 30 to 45 minutes
For more delicate tissues like the CNS tissue or brain, a longer drying time at lower temperature (e.g. [?]) is recommended to avoid splitting and cracking of the section due to excess heat.
37°C for 24 hours or longer
Another alternative to drying is by the use of [?].
adhesives
are not necessary for routine staining, provided that the slides are clean and free from grease.
Adhesives
they are essential for methods that require exposure of sections to acids and alkalis (especially ammoniacal silver solutions) during staining.
Adhesives
There are still certain instances when sections may float from the slide:
For urgent cryostat sections to be submitted for immunocytochemistry
For central nervous system tissues
For tissues containing blood clot
For tissues which have been decalcified
When sections are to be subjected to high temperatures
The most commonly use adhesive is .
Albumin
Albumin solution is prepared by mixing equal parts of [?], then filtered through coarse filter paper and a crystal of Thymol is added.
glycerin, distilled water and white of eggs
One disadvantage is that it retains some of the stain and gives a dirty background.
albumin
also a favorite adhesive; can be bought as a 0.1 % solution and further diluted (1 in 10 with distilled water) when ready to use.
Poly-L-lysine,
Sections are coated with this and allowed to dry. With time, the adhesive ability of this substance slowly loses its effectiveness. Therefore the coated slides should be used within a few days.
Poly-L-lysine
is a better section adhesive and coated slides can be stored for a long time.
Aminopropyltriethoxysilane (APES)
Slides are dipped in 2% APES in acetone drained then dipped in acetone, drained again and finally dipped in distilled water.
Aminopropyltriethoxysilane (APES)
It is invaluable in cytology particularly for cytosine preparation of proteinaceous or bloody material.
Aminopropyltriethoxysilane (APES)
is the most commonly used because it is very easy to make, is convenient, and is relatively inexpensive.
Mayer’s egg albumin
For celloidin sections, egg albumin is smeared on the slide. The section is then transferred from 95% alcohol bath to the slide, pressed flat on the slide with a smooth filter paper coated with thin celloidin mixture.
Mayer’s egg albumin
- dried, and stored in 70% alcohol until it is ready for staining.
Dried Albumin
Adding up to 30 ml of [?] to the water in a floating out bath and mixing it well is a most convenient alternative to direct coating of slides.
Gelatin (1%)
Coat the slides with the above mixture. Allow coated slides to dry at 37°C for one hour or overnight before use.
Gelatin-formaldehyde mixture
This aqueous detergent can be purchased as a 0.1% solution which is further diluted 1:10 with distilled water (final dilution to 0.01%) prior to use. Sections are coated with this and allowed to dry.
Poly-L-Lysin e
This is widely used as a section adhesive in immunohistochemistry.
Poly-L-Lysin e
must be used within a few days after they are prepared, since its effectiveness as an adhesive slowly decreases in time.
Poly-L-Lysin e
are very useful in cytology, particularly for cytospin preparations of proteinaceous or bloody material.
APES (3-aminopropylthriethoxysilane)
The slides are dipped in 2% [?] in acetone, drained, dipped in acetone, drained again, and finally dipped in distilled water. They are then placed upright in a rack and allowed to dry.
APES (3-aminopropylthriethoxysilane)
are better than poly-L-lysine coated slides because they can be stored for a long time without losing their adhesiveness.
APES (3-aminopropylthriethoxysilane)
bonds specimen, slide and coverslip together with a clear durable film.
mounting medium
is 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
It protects the stained section from getting scratched, to facilitate easy handling and storage of the slides, and to prevent bleaching or deterioration due to oxidation, thereby preserving the slides for permanent keeping
mounting medium
helps prevent the distortion of image during microscopic examination.
mounting medium
are often chosen for a specific refractive index (R.I.), which can enhance specimen details or make them invisible
mounting medium
Materials like [?] become totally invisible if immersed in a solution of the same refractive index.
glass
is 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
The mounting media must always have an [?] higher than the mounted sample to impart more
RI
The slide may then be incubated at [?] after mounting, to harden the medium.
37°C for 12-24 hours
Do not store mounted slides vertically for [?] if cured at room temperature.
2 days
will cause it to ooze out of the sides of the cover glass, and should be carefully wiped with a fine cloth moistened with xylene.
Excessive mounting medium
will dry up the section, causing shrinkage and cracking of the specimen.
Excessive blotting
if not removed, will mix with the mountant and form bubbles on the slide.
Excess xylene
may also cause improper setting of the coverslip or formation of bubbles on the section, which can be teased out by gently pressing on the cover glass with a pointed forceps or mounting needle.
Too little mounting medium
Setting may be hastened in a hot oven at
50°C for 2 hours.
should be chosen that will not fade the particular stains used
mounting medium
