FIXATIVES

Jenivie Morillo Jose

University of Santo Tomas
National Children’s Hospital
 FIXATIVES

SIMPLE COMPOUND

Aldehyde
Micro-Anatomical

Metallic
Cytological
Picric Acid

Acetic Acid

Acetone

Alcohol

Osmium Tetraoxide
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I.ALDEHYDE FIXATIVES

-for ROUTINE PARAFFIN SECTIONS, FROZEN

SECTIONS, ELECTRON MICROSCOPY
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A.FORMALDEHYDE

 -produced by the OXIDATION of METHYL

ALCOHOL
 -soluble in WATER to extent of 40% by weight
 -powerful REDUCING AGENT

 FIXATION TIME: 12-24 hrs.

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ADVANTAGES:
 CHEAP, readily available, easy to prepare,
relatively stable
 PENETRATE tissue well
 Doesn’t OVERHARDEN tissues
 Preserves FAT and MUCIN
 BEST FIXATIVE for NERVOUS TISSUE
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DISADVANTAGES:
 Causes ALLERGIC DERMATITIS, ALLERGIC
RHINITIS and SINUSITIS
 SHRINKAGE of tissues
 REDUCES BASOPHILIC, EOSINOPHILIC
staining of cells
 Forms abundant BROWN ARTIFACTS,
PIGMENTS and GRANULES
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REMOVAL OF FORMALIN PIGMENTS:

 KARDASEWITSCH’S METHOD

 LILIE’S METHOD

 PICRIC ACID METHOD

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B. GLUTARALDEHYDE

 -aldehyde M.W. 100

 -2 FORMALDEHYDE residues linked by

straight 3 CARBON CHAINS
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ADVANTAGES:

 Stable nature, effect on tissues, giving firmer

texture and color

 Preserves CELLULAR and PLASMA PROTEIN

better
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DISADVANTAGES:

 MORE expensive

 LESS stable
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C.PARAFORMALDEHYDE

 -polymer of formaldehyde

 -white powder
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ADVANTAGES:

 EXCELLENT FIXATIVE for ROUTINE

PARAFFIN SECTIONS

 For THIN and ULTRATHIN sections for

PLASTIC EMBEDDING
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DISADVANTAGES:

 Expensive

 UNSTABLE
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II. METALLIC FIXATIVES

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A.MERCURIC CHLORIDE

-frequently used in SATURATED SOLUTION

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ADVANTAGES:

 PENETRATES and HARDENS tissue rapidly

 Gives BETTER STAINING of NUCLEI and
CONNECTIVE TISSUE
 recommended for RENAL, FIBRIN and
CONNECTIVE TISSUE
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DISADVANTAGES:

 SHRINKAGE of cells
 LESS stable
 Corrodes metals except NICKEL
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B.CHROMATE FIXATIVES

 -STRONG OXIDIZING AGENT

 -shouldn’t combine with REDUCING AGENTS
like ALCOHOL and FORMALIN
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ADVANTAGES:

 Strong PROTEIN PRECIPITANT

 Preserves CARBOHYDRATES
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DISADVANTAGES:

 Forms a PRECIPITATE of INSOLUBLE

suboxide

 POOR for GLYCOGEN FIXATION

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Potassium Dichromate
 -STRONG FIXATIVE for LIPIDS

ADVANTAGES:
 Fixes but DOES NOT PRECIPITATE CYTOPLASMIC CONTENTS
 Preserves MITOCHONDRIA

DISADVANTAGE:

 Penetrates tissue SLOWLY

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C.LEAD FIXATIVES

-ADVANTAGES:

 mainly for MUCOPOLYSACCHARIDES

 precipitates PROTEIN
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DISADVANTAGES:

 Penetrates tissue SLOWLY

 Expensive
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III. PICRIC ACID FIXATIVES

 STRONG or SATURATED SOLUTION

 -2,4,6-trinitrophenol
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ADVANTAGES:

 Best fixative for GLYCOGEN

 Penetrates tissue RAPIDLY

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DISADVANTAGES:

 LYSES red blood cells

 SHRINKAGE of tissues
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IV. ACETIC ACID FIXATIVE

ADVANTAGES:

 Fixes and precipitates NUCLEOPROTEIN

 Precipitates CHROMOSOMES, CHROMATIN

MATERIALS
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DISADVANTAGES:

 SWELLING of tissues

 DESTROYS MITOCHONDRIA and GOLGI

ELEMENTS
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V. ACETONE

 -used only in ENZYME STUDIES

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ADVANTAGES:

 Recommended for PHOSPHATASES and

LIPASES studies

 For fixing BRAIN TISSUES for RABIES

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DISADVANTAGES:

 EVAPORATES rapidly

 DISSOLVES fats
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VI. ALCOHOL FIXATIVES

 for RAPID DENATURING and

PRECIPITATION of PROTEINS by destroying
HYDROGEN and other BOND
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ADVANTAGES:

 IDEAL for SMALLER tissue FRAGMENTS

 EXCELLENT for GLYCOGEN preservation

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DISADVANTAGES:

 LYSES red blood cells

 DISSOLVES LIPIDS and FATS

 POLARIZATION of GLYCOGEN
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VII. OSMIUM TETROXIDE FIXATIVES

 -osmic acid

 -pale yellow

 -dissolves in water

 -STRONG OXIDIZING AGENT

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ADVANTAGES:

 Fixes CONJUGATE FATS and LIPIDS

permanently

 EXCELLENT NUCLEAR staining

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DISADVANTAGES:

 VERY expensive

 SLOW fixing agent

COMPOUND FIXATIVES

I. MICRO-ANATOMICAL FIXATIVES
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10 % FORMOL SALINE
 -for NERVOUS system and GENERAL post
mortem materials

10% BUFFERED FORMALIN

 -for post mortem SURGICAL RESEARCH
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HEIDENHAIN’S SUSA SOLUTION

 -for biopsies of the SKIN

FORMOL SUBLIMATE
 -for ROUTINE post mortem materials
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FORMOL SALINE SUBLIMATE

 -for post mortem materials

ZENKER’S SOLUTION
 -for post mortem materials
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ZENKER’S FORMOL (HELLY’S FLUID)

 -for PITUITARY TISSUES and BONES

BOUIN’S SOLUTION
 -for EMBROYS
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II. CYTOLOGICAL FIXATIVE

A. NUCLEAR FIXATIVE
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FLEMMING’S FLUID
 -for nuclear structures

CARNOY’S FLUID
 -for CHROMOSOMES STUDY, LYMPH
NODES, GLYCOGEN
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BOUIN’S FLUID
 -for EMBRYOS and GLYCOGEN

NEWCOMER’S FLUID
 -for MUCOPOLYSACCHARIDES, NUCLEAR
PROTEIN and CHROMOSOMES
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B. CYTOPLASMIC FIXATIVE

FLEMMING’S FLUID(minus ACETIC ACID)

 -for CYTOPLASMIC STRUCTURES

CHAMPY’S FLUID
 -for MITOCHONDRIA, GOLGI ELEMENTS and
FATS
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REGAUD’S FLUID(MOLLER)
 - for MITOCHONDRIA and YOLK

ORTH’S FLUID
 -for EARLY DEGENERATIVE PROCESS and
TISSUE NECROSIS
Thank you 