MLT GlobeHisto & Cyto Techniques: Fixation, processing & staining – page 2
94 Histo & Cyto Techniques MCQs on Fixation, processing & staining with answers and explanations.
Small tubular structures such as a vas deferens or fallopian tube are best embedded:
Embedding tubes on end gives transverse sections showing the lumen and all wall layers. Longitudinal orientation may miss the lumen or show only part of the wall.
Fresh tissue for a frozen section is usually attached to the cryostat chuck using:
OCT compound freezes to a firm support around the tissue and attaches it to the chuck, and it dissolves in water during staining. Paraffin cannot be used with unfixed frozen tissue.
Positively charged or silane-coated slides are used for immunohistochemistry mainly to:
Harsh steps such as heating in retrieval buffer can detach sections, and charged or coated slides hold them firmly. Endogenous peroxidase is blocked with hydrogen peroxide, not by the slide type.
Before H&E staining of a paraffin section, the first step is to:
Aqueous dyes cannot penetrate paraffin, so sections are dewaxed in xylene and brought through alcohols to water. Acid alcohol is used later as a differentiator.
In the Verhoeff–van Gieson stain, elastic fibers appear:
Verhoeff's iron hematoxylin stains elastic fibers and nuclei black. The van Gieson counterstain then colors collagen red and muscle or cytoplasm yellow.
Bacteria in a tissue section need to be classified as Gram-positive or Gram-negative. The appropriate stain is:
Brown–Hopps and Brown–Brenn are Gram methods adapted for tissue sections. GMS is mainly for fungi and does not separate Gram reactions.
Mast cell granules stained with toluidine blue appear purple-red, while the background stains blue. This property is called:
Metachromasia is staining in a different color from the dye itself, caused by stacking of dye molecules on dense anionic groups such as heparin. Birefringence is a polarization property.
Tissue cassettes should be labeled with pencil or solvent-resistant ink mainly because:
Alcohols and xylene during processing can dissolve normal ink, risking loss of specimen identity. Pencil, solvent-resistant markers or printed labels keep the identification.
Picric acid must be stored moistened with water because, when dry, it:
Dry picric acid (and its metal picrates) is shock-sensitive and explosive, so it is kept wet and away from metal lids. Its yellow color and fixing ability are not the concern.
A drawback of adding alcohol pre-fixative to fluid specimens is that it:
Alcohol precipitates protein into a background film and hardens cells, which may shrink and adhere poorly. It actually preserves cells and inhibits bacteria.
Which fixative is commonly recommended for cell block preparation from fluids?
AAF fixes the pellet firmly while formalin keeps it compatible with paraffin processing and immunostaining. Plain alcohols tend to give fragile, shrunken pellets.
Carnoy's fixative is useful for bloody specimens because:
The acetic acid hemolyzes red cells, clearing the background, and the alcohol gives good nuclear fixation. Carnoy's is rapid but causes considerable shrinkage.
After a cell pellet is clotted with plasma and thromboplastin, the resulting cell block is next:
The clot is wrapped, fixed in formalin, placed in a cassette and processed to paraffin, allowing sections and immunostains. Other steps skip sectioning.
Spray fixatives should be applied from a distance of about:
Spraying too close freezes and disrupts cells or blows them off; spraying too far leaves parts unfixed and air-dried.
Why are aerosol spray fixatives avoided for heavily bloody smears?
Spray on bloody material makes red cells clump and dry unevenly, hiding diagnostic cells. Such smears are better wet-fixed or treated with a hemolyzing fixative.
Before fixation, a thick needle core biopsy is often laid flat on:
Filter paper keeps the core straight and prevents curling, aiding orientation in the block. Gauze absorbs and distorts tissue.
The minimum time for wet fixation of Pap smears in 95% ethanol is about:
About 15 minutes allows complete fixation. Smears can stay in fixative much longer, but shorter times give poor nuclear detail.
Denatured alcohol is suitable as a Pap fixative when used at:
It substitutes for 95% ethanol only at high concentration; dilute alcohol fixes poorly and gives pale, swollen nuclei.
An acceptable substitute for 95% ethanol in Pap fixation is:
Absolute methanol (and denatured alcohol) fixes comparably to 95% ethanol. Dilute propanol, acetone and water give poor nuclear preservation.
The ether–alcohol cytology fixative is made of:
The classic Papanicolaou fixative is equal parts diethyl ether and 95% ethanol. It is now largely abandoned because ether is highly flammable.