Fixative Information Sheets

Vocabulary practice flashcards covering simple aqueous fixatives, compound/combined fixatives, and non-aqueous fixatives, including their chemical compositions, tissue effects, staining applications, and safety concerns.

Acetic acid

A non-coagulant

Non-additive

Causes tissue swelling

Leaves tissue soft

Preserves DNA and nucleoproteins

Lyses red blood cells

Does not fix lipids or carbohydrates

PEL of 10ppm

Formaldehyde

A colorless gas

An additive

Non-coagulant fixative that

Hardens tissue

Penetrates rapidly but fixes slowly

Reacts with amines, amino acids, and nucleic acids to add an aldehyde complex forming methylene bridges.

10% Formalin solution

A fixative solution composed of $1\text{ part}$ $37\%$ formaldehyde and $9\text{ parts}$ water.

Methylene bridges

Cross bridges created during formaldehyde fixation when an added aldehyde complex reacts across amines, amino acids, and nucleic acids.

Glutaraldehyde

A dialdehyde

Additive

Non-coagulant

Over-hardens tissue

Fixes at the rate it penetrates

Preserves cellular ultrastructures for electron microscopy (EM)

Causes false positive results in PAS staining due to an unreacted aldehyde group.

Glyoxal

A dialdehyde fixative

Additive

Non-coagulant

A $40\%$ aqueous solution that must be buffered to pH 4.0

Lyses red blood cells

Prevents nuclear bubbling

Results in poor staining of eosinophils and Paneth cells.

Mercuric chloride

An additive

Coagulant

Reacts with sulfhydryl and amino groups to form protein crosslinks

Enhances tissue receptivity to dyes

Prevents tissue freezing

Causes mercury pigment

Poses extreme toxicity to the central nervous system

Osmium tetroxide

An additive, non-coagulant fixative ($OsO_4$) used for electron microscopy that turns cell membranes black and electron dense, renders lipids insoluble, penetrates only a few cell layers deep ($1\,mm$ section limit), and has a time-weighted average (TWA) of $0.02\,ppm$.

Picric acid

An additive, coagulant fixative ($C_6H_3N_3O_7$) that decalcifies, softens, and shrinks tissue, leaves DNA soluble, makes tissue receptive to acid dyes, and is an explosive hazard that must be kept moist.

Potassium dichromate

An additive, non-coagulant fixative ($K_2Cr_2O_7$) that reacts with carboxyl and hydroxyl groups, preserves lipids and mitochondria, dissolves DNA, acts as a coagulant (chromic acid) in an acid solution, and is a known carcinogen.

Zinc salts

An additive, coagulant fixative ($ZnSO_4$) that preserves immunoreactivity, provides good morphological detail, exhibits a slow penetration rate, and can cause pigment formation.

Orth solution

A compound fixative containing potassium dichromate, sodium sulfate, distilled water, and formaldehyde added before use; used in pheochromocytoma diagnosis by staining cytoplasmic chromaffin granules orange to brown.

Chromaffin granules

Chromate-loving granules located in the cytoplasm of the adrenal medulla that stain orange to brown when exposed to chromate fixatives such as Orth solution.

B-5 fixative

A coagulant, additive compound fixative made of mercuric chloride, anhydrous sodium acetate, and distilled water; provides clear nuclear detail on hematopoietic and lymphoreticular tissue, with mercury pigment treatable using iodine and sodium thiosulfate.

Helly fixative

An unstable, coagulant compound fixative composed of mercuric chloride, potassium chloride, sodium sulfate, distilled water, and formaldehyde (added before use) that preserves red blood cells via chrome and formalin, but is unsuitable for silver stains.

Zenker fixative

A stable, coagulant compound fixative containing mercuric chloride, potassium chloride, sodium sulfate, distilled water, and acetic acid; acts as a nuclear fixative, lyses red blood cells, dissolves iron, and decalcifies needle biopsies of bone marrow.

Bouin solution

A compound fixative composed of picric acid, $37\text{--}40\%$ formaldehyde, and glacial acetic acid; provides soft texture and clear nuclei for GI tract and endocrine biopsies, lyses red blood cells, dissolves iron and calcium, and must be washed and stored in $70\%$ alcohol.

Gendre fixative

An additive, non-coagulant compound fixative containing $95\%$ alcohol saturated with picric acid, $37\text{--}40\%$ formaldehyde, and glacial acetic acid; specifically preserves carbohydrates such as glycogen.

Hollande fixative

A compound fixative consisting of picric acid, $37\text{--}40\%$ formaldehyde, glacial acetic acid, copper acetate, and distilled water; decalcifies small bone specimens and uses cupric acetate to stabilize red blood cell membranes, eosinophil granules, and endocrine cells.

Zamboni fixative

A stable compound fixative formulated with paraformaldehyde, picric acid, and phosphate buffer; preserves morphologic characteristics for electron microscopy and allows secondary fixation with osmium tetroxide.

Acetone

A rapid-acting, non-additive, highly flammable fixative that causes extreme shrinkage, distortion, and overhardening; used on frozen tissues for cell surface antigen immunohistochemistry (IHC) and on brain tissue for rabies diagnosis.

Methyl alcohol

A non-additive, coagulant non-aqueous fixative primarily utilized for blood smears and touch preparations.

Ethyl alcohol

A non-additive, coagulant non-aqueous fixative that dissolves fat, overhardens tissue, and preserves pigments and water-soluble components such as glycogen and gout-derived urate crystals.

Carnoy fixative

A rapid-acting non-aqueous fixative composed of absolute alcohol, chloroform, and glacial acetic acid; preserves glycogen and nuclear detail, lyses red blood cells for cytology, and is toxic to the central nervous system, liver, and kidneys.

Clarke fixative

A coagulant, non-additive fixative prepared before use from absolute alcohol and glacial acetic acid; provides good microanatomical preservation and lyses red blood cells.