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1. Why do you think there are specimen acceptance criteria?
Specimen acceptance criteria allow the unequivocal identification of the patient. Appropriate scrutiny of specimens and associated request cards is a fundamental tenet of good histology.
2. What are the changes cells and tissue undergo when they are removed from the body?
Cells removed from the body undergo autolysis (self-destruction) and putrefaction (destruction by microorganisms)
3. What are the principles of fixation?
The principles of fixation are to maintain cells in as life-like a state as possible without artefacts, in a manner which allows subsequent laboratory testing.
4. What might the implications of a poorly fixed specimen for accurate diagnosis?
Poorly fixed specimens have poor nuclear detail and may undergo epitope loss making immunocytochemistry less valuable.
5. What does formalin actually fix?
Formalin fixes proteins. In aqueous solution it forms methylene glycol in a reversible reaction, and reacts with amine, guanidine, sulphydryl and amide groups, and with lipids.
6. What are the key hazards associated with handling formalin?
- Causes burns
- Toxic by inhalation, ingestion and through the skin
- Possible human carcinogen
- May cause allergic reactions.
7. What factors affect the speed of fixation?
Speed of fixation is affected by a number of factors, including:
- Specimen size
- Temperature
- Faulty batches of solution
- Incorrect pH
8. What methods are used for decalcification?
Agents used to decalcify tissues fall into two broad categories: acids and chelating agents. In-house acid solutions based on nitric, hydrochloric or formic acid are common, while commercial solutions are also available, mostly incorporating hydrochloric or formic acid.
9. How might the decalcification of tissues be speeded up? What automation could be used?
The speed of decalcification is increased by:
- a higher concentration of acid used in the decalcifying agent
- agitation of the tissue during decalcification
- changing the decalcifying agent at regular intervals so that the solution does is not saturated with calcium ions.
10. How is the end point of decalcification tested? What alternative methods are available? Which is the best and why?
There are three main ways of assessing the end point of decalcification:
- X-ray. This provides the most accurate assessment of decalcification, but it is expensive.
- Chemical end-point testing. This relies on the fact that insoluble calcium ammonium oxalate is produce from free calcium ions in the decalcifying fluid, which if cloudy indicates that decalcification is incomplete.
- Manual manipulation. This is the least specific of the three methods.