Stains for Pigments, Minerals, and Cytoplasmic Granules

What is the purpose of the Prussian blue stain?

To demonstrate ferric rion (3+) in hemosiderin

What is hemosiderin?

A loosely bound protein complex containing ferric iron that forms when there is too much iron in the body

What are sections stained with in the Prussian blue stain?

Potassium ferrocyanide and HCl

What is the mechanism of the Prussian blue stain?

Potassium ferrocyanide combines with ferric iron to form insoluble pigment prussian blue

What is the fixative for the Prussian blue stain? What should be avoided?

NBF or alcohol, strongly acidic fixatives should be avoided because they can dissolve some iron

What is the control for the Prussian blue stain?

Section containing ferric iron but not too much or background staining will occur

What is the counter stain for the Prussian blue stain?

NFR

What else is stained red from the NFR in the Prussian blue stain?

Nuclei and hemofuchsin

What stains blue from the Prussian blue reaction?

Hemosiderin ferric iron

What is the purpose of the Turnbull blue stain?

To stain for ferrous (2+) iron in tissues

What is the significance for ferrous iron in tissues?

Not normally stored in tissues due to toxicity

What is the working solution for the Turnbull blue reaction?

Potassium ferricyanide and HCl

What is the mechanism of the Turnbull blue reaction?

Ferrous iron reacts with potassium ferricyanide to form insoluble pigment Turnbull blue

What is the fixative for Turnbull blue reaction?

Alcohol or NBF

What is the Turnbull blue slides rinsed with before counterstaining? What is the counterstain?

Acetic acid, NFR

What stains blue with the Turnbull blue reaction?

Ferrous iron (2+)

Does hemosiderin stain with the Turnbull blue reaction?

No because ferric iron does not stain, only ferrous, so hemosiderin is unstained to yellow

What is the purpose of the Schmorl technique?

Demonstration of reducing substances in tissue sections

Which reducing substances are demonstrated with the Schmorl technqiue?

Melanin, argentaffin granules and formalin pigment

What is the mechanism of the Schmorl technique?

Reducing substances present in tissue reduce ferric iron to ferrous iron (3+ to 2+)

What does reduced iron react with in the Schmorl technique?

Ferrous iron reacts with potassium ferricyanide to form insoluble Turnbull blue pigment

What fixative and control is used for the Schmorl technique?

NBF, control containing melanin or argentaffin granules

What pH does the Schmorl technique work best at?

2.4, adjust with HCl as necessary

What can be used as counterstains to the Schmorl technique?

Mucicarmine, metanil yellow, or NFR

What is stained blue to green with the Schmorl technique?

Reducing substances (melanin, argentaffin granules, formalin pigment)

What is stained rose in the Schmorl technique if mucicarmine counterstain is used?

Goblet cells, mucin

What is the purpose of the Fontana Masson stain?

To demonstrate reducing substances such as melanin, argentaffin granules, and neurosecretory granules

What is the mechanism of the Fontana Masson stain?

Argentaffin tissue components bind silver and reduce it to visible form themselves

What is the reducing agent in the Fontana Masson stain?

None, this is an argentaffin mechanism

What is the fixative for Fontana Masson? What should be avoided?

NBF, alcohol should be avoided because it dissolves argentaffin granules

What is the control for Fontana Masson?

If staining for melanin use a skin control, if staining for argentaffin granules use small intestine or appendix

What is the working solution for Fontana Masson?

Silver nitrate and ammonium hydroxide (diamine silver)

What is the purpose of gold chloride and sodium thiosulfate in the Fontanna Masson stain?

Gold chloride tones, sodium thiosulfate removes unreduced silver

What is the counterstain used with Fontanna Masson to stain nuclei pink to red?

NFR

What is stained black by the Fontanna Masson stain?

Melanin, argentaffin granules, reducing substances

What are some differences to the microwave Fontanna Masson stain?

Faster, lots of DI rinses, same stain colors achieved

How to differentiate melanin after Fontana Masson stain?

Melanin is bleached away by potassium permanganate with oxalic acid

What is the purpose of the Grimelius stain?

To demonstrate argyrophilic granules in neurosecretory tumors

How to differentiate between argentaffin and argyrophilic granules?

Both Fontanna Masson and Grimelius stains need to be ran and compared side by side

What is the fixative and control for Grimelius stain?

NBF, positive argyrophilic tumor control or small intestine

What is the working solution for the Grimelius stain?

Silver nitrate, acetic acid, acetate buffer to 5.6

What is in the reducing solution for the Grimelius stain?

Hydroquinone, sodium sulfite

What counterstain is used with the Grimelius stain to stain nuclei red?

NFR

What stains brown to black with the Grimelius stain?

Argyrophilic and argentaffin granules

What is the background color in the Grimelous stain?

Pale yellow brown

What is the purpose of the Churukian Schenk method?

Demonstration of argyrophilic granules in neurosecretory tumors

What fixative is used for Churukian Schenk method?

NBF

What are slides rinsed in before working solution of Churukian Schenk method?

Citric acid acidified water

What is the working solution of the Churukian Schenk method?

Silver nitrate

What is the reducing solution of the Churukian Schenk method?

Hydroquinone, water, and sodium sulfite

What is the counterstain for Churukian Schenk method that stains nuclei red?

NFR

How many cycles does the non microwave Churukian Schenk method use?

2

How many cycles does the microwave Churukian Schenk method use?

3

What is in the microwave method that is not in the original Churukian Schenk?

Citric acid glycine

What do argentaffin and argyrophilic granules stain with the Churukian Schenk method?

Black, looks a bit lighter than other staining methods but not mentioned in text

What is the purpose of the Gomori methenamine silver stain?

To stain and demonstrate urates in tissue

What is the mechanism for the Gomori methenamine silver stain for urates?

The urates react with silver and are reduced to metallic form

What is the fixative for the Gomori methenamine silver stain for urates?

Absolute alcohol, any contact with water will dissolve urate crystals

What is the working solution for the Gomori methenamine silver stain?

Silver nitrate and sodium borate

What is the counterstain for the Gomori methenamine silver stain for urates?

Light green

What color do urate crystals stain with Gomori methenamine silver stain?

Black, background is green

What else may be demonstrated with the Gomori methenamine silver stain?

Calcium if the deposits are large enough to withstand dissolution in working silver solution

What is the purpose of the Hall bile stain?

To demonstrate bilirubin in tissue and to distinguish bile pigments from other tissue pigments, to aid in the diagnosis of obstructive jaundice

What is the mechanism of the Hall bile stain?

Bilirubin is oxidized by ferric chloride in trichloroacetic acid to form biliverdin which is a green color

What fixative and control are used for the Hall bile stain?

NBF, tissue containing bile

What is in the Fouchet reagent used in the Hall bile stain?

Trichloroacetic acid, water, and ferric chloride

What is a good counterstain for the Hall bile stain?

Van gieson to show red collagen and yellow cytoplasm and muscle against the green bile

What is the purpose of the von Kassa stain?

To demonstrate calcium in tissue

What type of stain is the von Kassa for calcium?

Indirect

What is the mechanism of the von Kassa stain for calcium?

Silver reacts with the anions of calcium salts (CO3-, PO4-)

What reduces silver to metallic form in the von Kassa reaction?

Bright light

What is the fixative and control for von Kassa?

Alcohol preferred but NBF is okay, section known to contain calcium for positive control

What is used as a counterstain for von Kassa?

NFR

What does calcium salts stain with von Kassa?

Black

What may remove calcium salts from tissue giving a false negative for von Kassa?

Alcoholic iodine

What is a problem with using sodium thiosulfate in the von Kassa reaction?

The silver carbonate salt formed from the carbonate (CO3-) group of a calcium salt is soluble in sodium thiosulfate and therefore will not be demonstrated leading to a false negative

What is the purpose of the Lindquist rhodanine method?

To demonstrate copper in tissues

What is the significance of copper in the liver?

Wilsons disease, a genetic disorder causing copper to be accumulated in the body

What is the mechanism for the rhodanine stain?

Rhodanine binds and demonstrates the protein which copper is bound to, high affinity for copper

How does the rhodanine copper stain compare to the old rubeic acid stain for copper?

More sensitive but less specific, may give false positives

What is the fixative and control used for the rhodanine copper stain?

NBF, control is a section containing copper fetal liver is excellent

What is the working solution for rhodanine copper stain?

Rhodanine and water

What is used as a counterstain to rhodanine copper to stain nuclei blue?

dilute Mayer hematoxylin and sodium borate

What should be kept in mind with the rhodanine copper stain?

If copper concentration is low fading may occur and copper can be confused with lipofuscin

What should be kept in mind for the rhodanine copper counterstain?

Mayer hematoxylin should be dilute, if too much is used it will mask the copper

What color is copper stained with the rhodanine method?

Bright red to red yellow

Is there a microwave method for rhodanine copper stain?

Yes, working solution contains sodium acetate formalin and aqueous mounting media must be used, results are pretty much the same