Appendix: Light Microscopy Stains

APPENDIX

Hematoxylin and Eosin (H&E)

Hematoxylin acts as a basic molecule and stains cellular regions rich in acidic macromolecules (DNA or RNA) a purplish blue color. It is the most common stain for demonstrating cell nuclei and cytoplasm rich in rough ER. Usually used as the contrasting “counterstain” with hematoxylin, eosin is an acidic stain that binds to basic macromolecules such as collagen and most cytoplasmic proteins, especially those of mitochondria. Eosin stains regions rich in such structures a pinkish red color. Tissue sections showing only structures with shades of purple and pink are most likely stained with H&E.

Pararosaniline–Toluidine Blue (PT)

This dye combination stains chromatin shades of purple and cytoplasm and collagen a lighter violet. These stains penetrate plastic sections more readily than H&E and are used in this atlas primarily with acrylic resin-embedded sections to provide better detail of cell and tissue structures. Toluidine blue is also commonly used for differential staining of cellular components, particularly cytoplasmic granules.

Mallory Trichrome

This procedure employs a combination of stains applied in series that results in nuclei staining purple; cytoplasm, keratin, and erythrocytes staining bright red or orange; and collagen staining bright or light blue. Mallory trichrome is particularly useful in demonstrating cells and small blood vessels of connective tissue. Similar stains, such as Masson trichrome and Gomori trichrome, yield comparable results except that collagen stains blue-green or green.

Picrosirius-Hematoxylin (PSH)

The dye sirius red in a solution of picric acid stains collagen red and cytoplasm a lighter violet or pink, with nuclei purple if first stained with hematoxylin. Under the polarizing microscope, collagen stained with picrosirius red is birefringent and can be detected specifically.

Periodic Acid-Schiff Reaction (PAS)

This histochemical procedure stains complex carbohydrate-containing cell components, which become magenta (shades of purplish pink). PAS is used most commonly to demonstrate cells filled with mucin granules, glycogen deposits, or the glycocalyx.

Wright-Giemsa Stain

These are two similar combinations of stains that are widely used on fixed cells of blood or bone marrow smears to demonstrate types of blood cells. Granules in leukocytes are seen to have differential affinity for the stain components. Nuclei stain purple and erythrocytes stain uniformly pink or pinkish orange.

Silver or Gold Stains

Various procedures employing solutions of silver or gold salts have been developed to demonstrate filamentous structures in neurons and fibers of reticulin (type III collagen). By these “metal impregnation” techniques these filaments stain dark brown or black. Such stains have been largely replaced now by immunohistochemical procedures.

Stains for Elastin

Several staining methods have been developed to distinguish elastic structures from collagen, most of ...