MLT GlobeHistopathology: Decalcification
14 Histopathology MCQs on Decalcification with answers and explanations.
Which acid is most widely used for routine decalcification in histology?
Formic acid is a relatively gentle yet effective decalcifier; strong mineral acids work faster but damage staining.
Which reagent is used to decalcify bone rather than to fix tissue?
Dilute nitric acid removes calcium salts and is a rapid decalcifier. Formalin, Bouin's and Carnoy's are fixatives.
Which acid, at around 5% concentration, is widely used as a rapid bone decalcifier?
5–10% nitric acid decalcifies quickly but can damage staining if overexposed. Formic acid is a gentler alternative; acetic and picric acids are fixative components.
At what stage should bone be decalcified?
Tissue must be fully fixed first; decalcifying unfixed tissue causes severe damage. Surface decalcification of a block is only an occasional supplement.
Formic acid used to decalcify a bone biopsy is usually prepared at about what strength?
Formic acid at around 5–10% decalcifies steadily while sparing staining; very strong acid damages tissue, very weak acid is too slow.
Which approach is standard today for processing a bone marrow trephine biopsy?
Current practice uses buffered formalin (or similar) and gentle decalcification to preserve morphology and antigens for immunohistochemistry.
Which acid releases only one hydrogen ion per molecule (monobasic)?
HNO3 has one replaceable hydrogen; sulphuric, oxalic and succinic acids each have two.
Which acid is favoured as a gentler decalcifier than nitric acid, giving better preservation of staining and antigens?
Formic acid (5–10%) decalcifies more slowly but damages tissue less than strong mineral acids, so it suits routine bone and marrow work. EDTA is gentler still.
Which decalcifying agent works by chelation and best preserves DNA and antigens, although it is slow?
EDTA binds calcium ions at neutral pH, so it spares nucleic acids and epitopes needed for IHC and molecular tests. Acids are faster but more damaging.
Which procedure will NOT make bone suitable for light microscopic sections?
Alcohol does not remove mineral, so bone stays too hard to cut on a standard microtome. Acid or EDTA decalcification, or resin embedding with hard knives, enables sectioning.
When EDTA is used for decalcification, the solution is adjusted to about which pH?
EDTA chelates calcium best near neutral pH (about 7–7.4), which is gentle and preserves antigens and DNA. It is slow compared with acid decalcifiers.
A small bone fragment must be fixed and decalcified at the same time. Which fixative can do both?
Zenker's contains acetic acid, which slowly decalcifies small specimens during fixation. NBF, glutaraldehyde and Zamboni lack sufficient acid.
Bone left too long in a strong acid decalcifier will most likely show:
Over-decalcification damages nucleic acids so nuclei stain poorly with haematoxylin. Cracking usually reflects overheated or overdried sections.
In electrolytic decalcification, how is calcium removed from bone?
An acid bath (typically formic plus hydrochloric acid) is used with a direct current, which attracts Ca²⁺ ions to the cathode and speeds decalcification. EDTA chelation is a separate, slower method.