Part 2 Flashcards
(52 cards)
Tissue blocks should be in what size?
Small (1 to 2mm^2 for EM and 2cm^2 for LM)
T/F: Large solid tissue, such as uterus, should be opened or sliced thinly to improve penetration of fixatives
TRUE
Contents that can inhibit the penetration of fixative, as well as damage tissue during sectioning, and therefore must be removed before fixation.
Fecal matter and stomach contents
Brain is usually suspended whole in
10% buffered formalin for 2-3 weeks to ensure fixation and some hardening prior to sectioning
T/F: The brain must be fixed before “grossing” or sectioning.
TRUE
Fixative that penetrate the best and worst
Best = Formalin and alcohol
Worst = Glutaraldehyde
To maintain an adequate fixation time of
4-6 hours
Recommended size of tissue
2cm^2 and no more than 4mm thick
The best results are usually obtained using
Slightly hypertonic solutions (400-450mOsm; isotonic solutions are 340 mOsm)
Commonly added to osmium tetroxide fixatives for electron microscopy.
Sucrose
Normally used as a 10% solution
Formalin
The presence of a buffer causes
Polymerization of the aldehyde, with consequent decrease in its effective concentration
Found to be an ideal concentration for immuno-electron microscopy
Low concentration of glutaraldehyde (0.25%)
Primary fixation in buffered formalin is usually carried out for
2-6 hours during the day when the specimen is obtained
Most of the formalin can be washed out after fixation for
24 hours
Prolonged fixation may cause
Shrinkage and hardening of tissue, and may severely inhibit enzyme activity and immunological reactions,
Remedy to restore the activity of some enzymes
Washing of tissue
For electron microscopy, it is recommended that diced tissues be fixed for
3 hours then placed in a holding buffer
Artefacts will be introduced by drying, so if the tissue is left out in the open, it should be kept moist with
saline
May act as mordants or accentuators to promote and hasten staining, or they may inhibit certain dyes in favor of another
Formaldehyde intensifies while osmium tetroxide inhibits
T/F: Fixatives increase the optical differentiation of cells and tissue components thereby rendering them more readily visible during examination.
TRUE
Characteristics of a GOOD Fixatives (9)
- It must be cheap.
- It must be stable.
- It must be safe to handle.
- It must kill the cell quickly thereby producing minimum distortion of cell constituents.
- It must inhibit bacterial decomposition and autolysis.
- It must produce minimum shrinkage of tissue.
- It must permit rapid and even penetration of tissues.
- It must harden tissues thereby making the cutting of sections easier.
- It must be isotonic, causing minimal physical and chemical alteration of the cells and their constituents
- It must make cellular components insoluble to hypotonic solutions and render them insensitive to subsequent processing.
- It must permit the subsequent application of many staining procedures to facilitate easier and more profitable examination.
Protein-denaturing agents.
Alcohol based fixatives
Act by forming insoluble metallic precipitates like mercuric chloride and picric acid
Metallic Fixatives
