HISTOPATH: OTHER STAINING SOLUTIONS

Van Gieson Stain

• Acid fuchsin and picric acid

• Demonstrate connective tissues

• Acid fuchsin (Masson stain)

◦ Stain collagen, smooth muscle, mitochondria

Acridine Orange

• Identify dead and living cells

• Green fluorescence - DNA

• Red fluorescence -RNA

• Nucleic acid selective fluorescent cationic dye useful for cell

Acridine Red 3B

• Demonstrate deposits of calcium salts and sites of phosphatase activity

Alcian Blue

• Same as chlorophyll

• Stains acid mucopolysaccharides

• Excellent stain

• Resistant to counterstains

• Specific for connective tissues and epithelial mucin

Aniline Blue

• Cytoplasmic stain

• Counterstain epithelial sections

• Aniline blue + distilled water + glacial acetic acid

Basic Fuchsin

• Plasma stain

• Deep staining of acid fast organisms

• Mitochondria

• Differentiation of smooth muscles

• Main ingredient of Feulgen’s and Schiff’s reagent: detect aldehydes

• Van Gieson: connective, mucins, elastic tissue

Benzidine

• Stain hemoglobin

Bismark Brown

• Contrast stain for

◦ Gram’s technique

◦ Acid fast

◦ Papanicolaou method

• Stain diphtheria organisms

Carmine

• Chromatin stain for fresh materials

• Combined with aluminum chloride to stain glycogen (Best carmine solution)

Celestine Blue

• Oxazine dye, alternative to iron hematoxylin nuclear stain

• Resistant to strong acid dyes

• Routine stain for fixed tissues

• Good nuclear definition

Congo Red

• Indicator

• Stain for axis cylinders in embryos

• Used as 4% aqueous solution in Krajan’s method — elastic tissues, amyloid & myelin

Crystal violet

• Nuclear stain

• Stain amyloid in frozen sections, platelets

• Gentian violet: crystal violet + methyl violet + dexterin

Cresyl violet

• Stain nervous tissues

• Stains neuronal cytoplasm (Nissl bodies)

Giemsa

• Mixture of methylene blue and eosin

• Stain blood to differentiate leukocytes

• Mostly used in methanol fixed blood films where it stained erythrocytes, pink, and the different types of leukocytes, allowing their identification according to size and shape of their nucleus

Gold Sublimate

• Stain used for metallic impregnation

• Gold chloride + mercuric chloride

Iodine

• Oldest stain

• Used for microscopic study of starch granules

• Stains amyloid, cellulose, starch, carotene & glycogen; useful in histological and biochemical applications

• Used for removal of mercuric fixative

artifact pigments

• Reagent to alter crystal and methyl violet
◦ Gram’s Iodine - identify and

differentiate bacteria

◦ Lugol’s Iodine - turns black in the

presence of starches

Gram’s Iodine

identify and differentiate bacteria

Lugol’s Iodine

turns black in the presence of starches

Janus Green B

• Demonstrate mitochondria during intravital staining

Malachite Green

• Contrast stain for ascaris eggs and erythrocytes

• Bacterial spore stain

• Used as decolorizer and counterstain

Methyl Green

• Stains chromatin green

• Gives false positive reactions - mucin

• Used commonly with brightfield microscope to dye the chromatin of cells for easy viewing

Methylene Blue

• Basic nuclear stain employed with eosin; to provide marked differentiation of various structures in the tissue

• Methylene blue stains acidic cell parts like nucleus and is a good counter stain with Eosin Y

• “Polychroming” - oxidation of methylene

blue: loss of methyl groups and leaving lower

homologues of the dye and deaminized

oxidation products

• Stain for plasma cells

• Cytological examinations of fresh sputum

for malignant cells

• Bacterial stain

• For diagnosis of diphtheria

• Vital staining of nervous tissue

Nile Red

• also known as Nile blue oxazone

• Formed by boiling Nile blue with sulfuric acid

• Lipophilic stain, it will also accumulate in lipid globules inside cells

Oil Red O

• Stain for neutral lipids and fatty acids

• Rapid and simple stain

• Useful in identifying fat emboli in lung or clot sections in peripheral blood

Orcein

• Excellent stain for elastic fibers

• Recommended for dermatological studies: due to the ability to demonstrate the finest and most delicate fibers in the skin

Osmium Tetroxide

• Selective stain for unsaturated lipids and lipoproteins like myelin → black

• Useful in lipid histochemistry and electron microscopy

Periodic Acid Schiff

• Stains glycogen, mucin, mucoproteins, glycoproteins, basement membranes, capsules, blood vessels, fungi, and intracellular carbohydrates

• Strongly stains mucus-secreting cells

• Glycogen staining can be removed with an amylase pre-digestion step

• Works by periodic acid oxidation of hydroxyl groups into aldehydes, which react with Schiff reagent to produce a reddish-purple color

Prussian Blue

• Colored salt of ferric ferrocyanide

• Used for microanatomical color contrast of specimens

• Demonstration of blood and lymph vessel by injection or intravital staining

Rhodamine B

• Used with osmic acid to fix

• Stain blood and glandular tissues

Silver Nitrate

• Used in 10% aqueous solution: to prepare various dilutions to be used in identification for spirochetes, reticulum and other fiber stains

Safranin O

• Nuclear stain

• Primarily as counterstain in Gram staining

• Yellow color → collagen

Toluidine Blue

• Nuclear stain for fixed tissues

• Substitute for thionine in frozen sections

• Staining Nissl granules or chromaphilic bodies

• Versatile dye that stains nuclei blue and can be used to differentiate different types of granules including mast cells

Victoria Blue

• Demonstration of neuroglia in frozen sections

Wright stain

• For blood cells

• 4 major staining properties:

◦ Basophilia (affinity for methylene blue):

stains basophils and nucleic acids

◦ Azurophilia (affinity for oxidation of

methylene blue): stains azurophilic granules

◦ Acidophilia (affinity for eosin): stains

eosinophilic structures e.g RBCs and

eosinophil granules

◦ Neutrophilia (affinity for complex of

dyes): stains neutrophils

4 major staining properties of WRIGHT STAIN

Basophilia

Azurophilia

Acidophilia

Neutrophilia

Basophilia

• stains basophils and nucleic acids

• affinity for methylene blue

Azurophilia

• affinity for oxidation of methylene blue

• stains azurophilic granules

Acidophilia

• affinity for eosin

• stains eosinophilic structures e.g RBCs and eosinophil granules

Neutrophilia

• affinity for complex of dyes

• stains neutrophils