Flashcards
Histological Slide Preparation
Cards in this deck (21)
- 1list the steps of histological preparation in orderfixation
dehydration
embedding
sectioning
staining
mounting - 2why must tissues be fixedso they don't rot
- 3what does fixation involve?denaturation of proteins
- 4common fixation methodschemical fixation
heating
cooling/freezing - 5what is the common method of fixation for light microscopy?chemical fixation
- 6what are the most frequently used chemical fixatives?aldehydes
- 7what chemical fixing agent is used in light microscopy and what is used for electron microscopy? why are they used for different types of microscopy?formaldehyde
glutaraldehyde - penetrates faster and more thoroughly than formaldehyde but is more expensive and toxic - 8outline the dehydration processtissue is immersed in a series of progressively stronger alcohols (50, 70, 90, 100%)
next alcohol is dispersed by immersing in a solvent such as xylene or cedarwood oil - 9why must tissue be embedded?after dehydration, the tissue is too soft to allow it to be cut into fine sections - must be embedded in something hard
- 10what is the most commonly used embedding agent?paraffin wax
- 11state the other embedding agents used in electron microscopyplastics based on methacrylate
- 12what is the embedded block of tissue mounted on for sectioning?microtome
- 13what is the thickness of the tissue sections for light microscopy? what about electron microscopy?7-10 microns
less than 2 microns - 14why are sections thicker than 12 microns harder to interpret in light microscopy?cellular components become superimposed
- 15how are sections made to adhere to the glass for preparation for staining?gentle heating of the slide until the wax melts
- 16tissue must be stained. stains are aqueous, what must happen before this can occur?the specimen was dehydrated and now has to be rehydrated.
sections are immersed in progressively weaker alcohols (100, 90, 70, 50%, distilled water)
the specimen is now ready to be stained, usually by immersion - 17how does this stain: Haematoxylin and Eosin H&E colour the specimen?basic components coloured pink/red (E)
acidic components coloured blue/purple (H) - 18what stain stains nuclei lighter than cytoplasmmethylene blue
- 19what does mounting entail?placing the specimen in a refractive medium
such media are not water based and so the specimen has to be dehydrated by immersion in progressively stronger alcohols
then covered with a drop of refractive mountant
finally a coverslip is applied and the preparation is allowed to dry - 20outline what stress may affect what is seen when viewing a stained sectionmechanical trauma - artificial folds, tears, spaces
poor quality/preservation of tissue - inadequate fixation, embedding, staining
water droplets confuse view - dehydration process incomplete
poor quality sections should be chucked out (stain badly) - 21what are some of the problems with interpreting histological samples?folding
artefactual spaces
air bubbles
water particles
interpretation of 3D structure from 2D image
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- FY Histology Flashcards on Histological Slide Preparation , created by Erica Lai on 16/11/2017.
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- Erica Lai
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