MLT GlobeHistopathology: Fixation & fixatives – page 2
50 Histopathology MCQs on Fixation & fixatives with answers and explanations.
Which fixative component lyses red blood cells?
Acetic acid lyses erythrocytes, so it is avoided when RBCs must be preserved; formaldehyde preserves red cells.
A recognised drawback of coagulant fixatives such as ethanol is that they:
Coagulants precipitate proteins, shrinking and hardening tissue; methylene bridge cross-linking is how formaldehyde (non-coagulant) acts.
B-5 fixative was once favoured for lymph node biopsies. Why is it now largely abandoned?
B-5 gave excellent nuclear detail but its mercury content is hazardous; neutral buffered or zinc formalin are now used.
Which factor most slows formalin fixation of a large, bulky specimen?
Formalin penetrates roughly 1 mm per hour, so thick specimens fix poorly in the centre unless sliced thinly.
Brown-black granules of formalin pigment are seen in a bloody tissue section. Which reagent removes them before staining?
Acid formaldehyde haematin is removed with alcoholic picric acid or alcoholic ammonia; buffered formalin prevents it forming.
H&E sections of a resected small bowel show loss of much surface epithelium, though deeper layers look normal. The most likely cause is:
Gut epithelium autolyses within minutes without fixation; true ulceration would show inflammation and granulation tissue.
Picric acid is a main ingredient of which fixative?
Bouin's contains picric acid, formaldehyde and acetic acid; picric acid is also used as a yellow dye in van Gieson.
Which fixative is traditionally recommended when lipids are to be demonstrated histochemically?
Calcium helps preserve phospholipids and formalin does not dissolve lipids; alcoholic fixatives extract lipids.
Commercial concentrated 'formalin' (labelled 100%) contains about how much formaldehyde?
Stock formalin is about 37–40% formaldehyde gas in water; '10% formalin' is therefore roughly 4% formaldehyde.
Why is 10% neutral buffered formalin preferred over Zenker's fluid for routine surgical specimens?
Zenker's contains mercuric chloride, which is toxic and leaves pigment; NBF is safer, cheaper and compatible with IHC and molecular tests.
A pathologist wants especially bright, crisp cytoplasmic and trichrome colours. Which traditional fixative is known for this effect?
Mercuric chloride fixatives such as Zenker's give brilliant cytoplasmic staining (mercury pigment must be removed first). Formalin gives adequate but duller colours; osmium tetroxide blackens lipids.
Cold acetone is often chosen to fix frozen sections for enzyme histochemistry and immunofluorescence because it:
Acetone precipitates proteins without cross-linking, so many enzymes and epitopes remain reactive. It is a poor morphological fixative and is not used for routine paraffin work.
Gastric washings sent for cytology or mycobacterial culture should be neutralized promptly because their acidity damages cells and organisms. A suitable neutralizer is:
Gastric acid rapidly destroys cells and reduces mycobacterial viability, so washings are neutralized with dilute NaOH (or sodium bicarbonate). Adding acid or peroxide would worsen damage.
Formalin for fixation is buffered to neutral pH mainly to prevent which artefact?
Acidic formalin reacts with hemoglobin to deposit birefringent brown pigment in blood-rich tissue. Buffering to pH ~7 prevents this; mercury pigment arises from mercuric fixatives.
Working 10% neutral buffered formalin contains approximately what concentration of formaldehyde?
It is a 1 in 10 dilution of 37–40% stock formaldehyde, so it contains about 4% formaldehyde. The '10%' refers to the dilution of formalin.
Why must tissue be well fixed before it is placed in alcohol for dehydration?
Fixation stabilizes proteins so the tissue withstands dehydration, clearing and hot wax. Unfixed tissue shrinks, hardens and distorts in graded alcohols.
Which fixative acts mainly by denaturing and coagulating proteins rather than cross-linking them?
Alcohols are coagulant fixatives that dehydrate and precipitate proteins. Formaldehyde and glutaraldehyde are cross-linking (additive) fixatives.
In formalin-fixed nuclei, what material commonly clumps into strands separated by clear spaces?
Formalin fixation causes nuclear chromatin to coalesce into strands with intervening clear areas. This is a nuclear, not cytoplasmic, change.
A brown-black granular deposit seen in blood-rich, formalin-fixed tissue shines when viewed between crossed polarizers. It is most likely:
Formalin pigment forms from haemoglobin in acidic formalin and is birefringent. Melanin, lipofuscin and bile pigment are not birefringent.
Which reagent is used in histology both as a fixative component and as a dye in its own right?
Picric acid fixes (in Bouin's) and is also a yellow acid dye used as a counterstain, for example in Van Gieson. Bouin's is a mixture, not a single substance.