HT Routine & Special Staining Methods 3 — Questions and Answers
Question 1: In the Alcian blue stain performed at pH 2.5, which tissue component is demonstrated?
- Acidic mucopolysaccharides (acid mucins) (Correct answer)
- Neutral mucins
- Basement membranes
- Nucleic acids
Correct answer: Acidic mucopolysaccharides (acid mucins)
Alcian blue at pH 2.5 stains acidic mucopolysaccharides blue by ionic bonding to negatively charged sulfate and carboxylate groups.
Question 2: Which stain best demonstrates reticular fibers (type III collagen) in lymph node architecture?
- Gordon and Sweet's reticulin silver stain (Correct answer)
- Masson trichrome
- Verhoeff's elastic stain
- Movat pentachrome
Correct answer: Gordon and Sweet's reticulin silver stain
Gordon and Sweet's silver impregnation method selectively coats reticular fibers black, making them distinct from other connective tissue elements.
Question 3: The Luxol fast blue (LFB) stain is used in neuropathology to demonstrate:
- Myelin sheaths (Correct answer)
- Axons
- Astrocytes
- Microglia
Correct answer: Myelin sheaths
Luxol fast blue binds to lipoproteins in myelin sheaths, staining them blue-green, and is often combined with cresyl violet or H&E for counterstaining.
Question 4: When performing the Giemsa stain on bone marrow, what is the purpose of the methanol fixation step?
- Fix cells to the slide and prevent washing off during staining (Correct answer)
- Decolorize background erythrocytes
- Enhance nuclear detail by hydrolyzing DNA
- Mordant the tissue for Romanowsky dyes
Correct answer: Fix cells to the slide and prevent washing off during staining
Methanol fixation adheres cells to the glass slide and preserves cellular morphology before the aqueous Giemsa solution is applied.
Question 5: In the von Kossa stain, calcium is demonstrated indirectly because the stain actually deposits:
- Silver on phosphate or carbonate anions associated with calcium (Correct answer)
- Calcium itself as a black precipitate
- Iron-calcium complexes
- Alizarin red chelated to calcium ions
Correct answer: Silver on phosphate or carbonate anions associated with calcium
Von Kossa silver nitrate is reduced in the presence of phosphate and carbonate salts co-located with calcium, depositing metallic silver that appears black.
Question 6: Which reagent in the Gram stain is responsible for trapping crystal violet-iodine complexes inside gram-positive cell walls?
- Iodine (mordant) (Correct answer)
- Safranin (counterstain)
- Acetone-alcohol (decolorizer)
- Crystal violet (primary stain)
Correct answer: Iodine (mordant)
Iodine acts as a mordant, forming large crystal violet-iodine complexes that become trapped within the thick peptidoglycan layer of gram-positive organisms.
Question 7: The Hall's bile pigment stain uses which reagent to convert bilirubin to biliverdin, which then stains green?
- Fouchet's reagent (ferric chloride in trichloroacetic acid) (Correct answer)
- Lugol's iodine
- Potassium ferrocyanide
- Sodium nitroprusside
Correct answer: Fouchet's reagent (ferric chloride in trichloroacetic acid)
Fouchet's reagent oxidizes bilirubin to biliverdin, which appears green, while connective tissue is counterstained with van Gieson solution.
HT Routine & Special Staining Methods 3
In the Alcian blue stain performed at pH 2.5, which tissue component is demonstrated?