ALDEHYDE FIXATIVES

FIXATIVE NOTE USE FIXATION ADVANTAGE/S DISADVANTAGES PRECAUTION/S

TIME
5% -Anti-fungal -Routine paraffin -24hrs -Cheap -Irritation -Prolonged storage at
FORMALDEHYDE property -Most sections -Electron -Readily available -Produce considerable low temperature ->
OR common fixative Microscopy -Easy to prepare shrinkage of tissues white ppt. -For fatty
10% -Histochemical and -Does not require washing out - -Prolonged period of tissues, add
FORMALDEHYDE Enzyme studies Does not over harden the tissue fixation bleaches specimen cadmium or cobalt salts
even with prolonged period of UNBUFFERED -Add magnesium carbonate
fixation -formalin may form abundant & calcium carbonate to
brown pigment granules (on neutralize acid reaction
blood containing tissue)
-reduces basophilic &
eosinophilic of cell
10% FORMOL -NaCl + Distilled H2O -Recommended for -24hrs @ -Allows natural tissue colour to -Similar with
SALINE -Simple CNS -postmortem 35’C be restored formaldehyde -Slow
microanatomic tissues for -48hrs @ -Ideal for most staining fixative
fixative histochemical exam 20’C – techniques including Silver -Metachromatic reaction with
-Preserves Fats, 25’C Impregnation amyloid is reduce
Enzymes, Mucin, -Large specimen may be fixed
Nuclear Protein for a long period of time
10% NEUTRAL -Best fixative -Surgical, Post mortem 4-24 hrs -Prevent precipitation of -Longer to prepare (time
BUFFERED (GENERAL) & Research specimen formalin -Fast penetration consuming)
FORMALIN -Fe pigments & -Hardens tissue better
Elastic fibers
-Immunohistchemistry
& Molecular tests
-Preserves
Fats,Mucin,
Glycogen

FORMOL- -A formal-mercuric -Recommended for 3-4hrs -Penetrates small pieces of tissues -Slow penetration
CORROSIVE chloride solution routine post mortem rapidly -Form a mercuric
(FORMOL- tissue -Brightens cytoplasmic and chloride deposits
SUBLIMATE) metachromic stains better than -Inhibits determination of the
formalin alone extent of tissue
-Excellent for silver decalcification
reticulum methods

GLUTARALDEHYDE -Enzyme 2-4hrs -Stable effect -More expensive

histochemistry - (2.5% sol’n) -Less tissue shrinkage -Reduces reactivity of mucin
Electron Microscopy -Less irritating (does not cause to PAS***
6 to 24hrs dermatitis)
(4% sol’n) -More satisfactory on tissue
giving firmer texture & better
tissues section

METALLIC FIXATIVES
FIXATIVE NOTE USE FIXATION ADVANTAGE/S DISADVANTAGES PRECAUTION/S

TIME
MERCURIC -Most common -For tissue photography 24hrs -Excellent for trichrome -Causes shrinkage of -Avoid using metal
CHLORIDE metallic fixative -Preserves renal tissues, staining -Best for cells -Remove iron from caps, jewelries
fibrin, CT, muscles metachromatic staining
FORMULA hemosiderin
Mercuric-chloride -Inert to fats and lipids
Potassium- -Formation of black
dichromate Na granular deposits
sulphate -Reduce demonstration
Distilled H2O of glycogen
 ZENKER’S FLUID FORMULA -Recommended of 12 – 24hrs -Gives excellent staining -Unstable after addition -Wash tissues thoroughly in
HgCl – 5g trichrome staining result -Stable of Acetic acid running water
K dichromate – 2.5g -Recommended for -Form mercuric chloride
Na sulphate – 1g fixing small pieces of precipitates
Distilled H2O – liver, -Does not permit cutting
100mL Glacial acetic spleen, connective of frozen sections
acid – 5mL tissue fibers and nuclei.

ZENKER-FORMOL FORMULA -Recommended for 12 - 24hrs -Penetrates & fixes tissue -Form mercuric chloride
(HELLY’S SOLUTION) HgCl – 5g fixing pituitary gland, well -Nuclear fixation better deposits
K dichromate – 2.5g Bone than Zenker’s fluid
Na sulphate – 1g marrow, Spleen, Liver -Preserves cytoplasmic granules
Distilled H2O – well
100mL Strong
formaldehyde – 5mL
HEIDENHAIN’S Made up of MgCl2, -Tumor biopsy 3 – 12hrs -Penetrates & fixes tissues -Prolonged fixation -After fixation, tissue should
SUSA SOLUTION glacial acetic acid -Excellent cytologic rapidly -Permits most staining cause shrinkage be transferred directly to
and formalin fixative procedures hardening and high grade alcohol to avoid
-Easy sectioning of large blocks bleaching swelling
of fibrous connective tissue -Weigert’s method is not
-Reduces processing time by possible
transferring directly to 95% or - Form mercuric
absolute alcohol chloride precipitates

Prepared by: ROBIN C. AT-AT, RMT, MPH

SCHAUDINN’S FORMULA -For wet smear 30mins – -Very good cytoplasmic fixative -Caused excessive -Wash tissues thoroughly in
FIXATIVE Mercuric Chloride preparation and small 12hrs shrinkage - Form mercuric running water
(unsaturated) – 66mL connective tissues chloride precipitates - After fixation, tissue
95% Ethanol – 33mL -Qualitative fixation should be transferred
Glacial acetic acid – of protozoan directly to 70% alcohol
1mL before staining
B-5 FIXATIVE FORMULA - Commonly used for 2 – 4hrs -Carcinogenic & Irritant
Mercuric chloride – BM biopsy and some - Form mercuric
12g Na acetate – 2.5g lymph nodes when chloride precipitates
Distilled H2O – 200mL lymphomas are
suspected

CHROMATE FIXATIVES
FIXATIVE NOTE USE FIXATION ADVANTAGE/S DISADVANTAGES PRECAUTION/S

TIME
CHROMIC ACID -Fixes carbohydrates -Strong oxidizing agent -Formaldehyde must be added
-Precipitate all CHON before used to counteract
effects & prevent
decomposition of
solution upon long
standing
POTASSIUM pH 4.5 – pH 5.2 -Preserves lipids and -Fixes but does not precipitates -If solution is acidified, -Acidify solution to fix
DICHROMATE mitochondria cytoplasmic structures mitochondria is destroyed Cytoplasm, Chromatin
(K2Cr2O7) bodies and Chromosomes
REGAUD’S -Recommended for 12 – 48hrs -Penetrates tissue well -Slow penetration
(MOLLER) FLUID demonstration of Golgi -Hardens tissue better & more -Form precipitates of sub
bodies, Mitochondria, rapidly than ORTH’s oxides
RBC, Chromatin, Colloid -Blacken tissue pigments
-Poor nuclear standing
containing tissue and
Mitotic figures
ORTH’S FLUID -Study of early 36 – 72hrs -Preserve myelin better than - Slow penetration
degenerative processes buffered formalin -Form precipitates of sub
& tissue necrosis oxides
-Blacken tissue pigments
-Poor nuclear standing

PICRIC ACID FIXATIVES

 FIXATIVE NOTE USE FIXATION ADVANTAGE/S DISADVANTAGES PRECAUTION/S

TIME
PICRIC ACID -Strong -Preserves glycogen 12 – 24hrs -Penetrates tissue well & -Reduce iron in tissue -Never wash picric acid-
saturated fixes small tissues rapidly -Causes frozen section to fixed tissues in water before
aqueous sol’n -Allows brilliant staining crumble when cutting
-Yellow stain with trichrome method -Picrate are formed dehydration
taken in by tissue -Precipitates all protein -Excessive staining will
-Stable cause yellowing of the
prevents small -Suitable for aniline stains tissue
fragments from (Mallory, Heidenhains, Masson)
being overlooked
BOUIN’S SOLUTION -Fixation of embryo 6 – 24hrs -Minimal distortion of -Soluble in water
micro anatomic structures -Not suitable for kidney
-Does not need washing structures
-Brilliant staining of tissues -Destroy cytoplasmic
structures
-Hemolysis & removes
ferric ion from blood
pigments
-Reduce faulgen reaction
BRASIL’S -Excellent fixative for glycogen 24hrs Better & less messy than
ALCOHOLIC bouin’s solution
PICROFORMOL

ALCOHOL FIXATIVE
FIXATIVE NOTE USE FIXATION ADVANTAGE/S DISADVANTAGES PRECAUTION/S

TIME
70% - 100% -Preserves nuclear stain 24hrs -Excellent for glycogen -Cause hemolysis
ALCOHOL -Causes glycogen granules to
move towards pole or ends
of the cells
100% METHYL -Dry and Wet smears 24 – 48hrs -Fixes and dehydrates at the -Slow penetration
ALCOHOL -Blood smears, Sputum same time -Over hardened & difficult
-Urine to cut (>48hrs)
-Bone marrow
70% - 100% 18 – 24hrs -Preserves but not fixes -Polarization of glycogen -Should not be mixed
ETHYL glycogen -Fixes blood, tissue granules with strong oxidizing
ALCOHOL films and smears -Hardening & shrinkage agent
-Fixed tissue pigments fairly well of tissues
-Hemosiderin preservation
less than buffered
formaldehyde
-Strong reducing agent
CARNOY’S FLUID -Rapid fixative -Chromosomes, Lymph 1 – 3hrs -Permits good nuclear staining -Slow penetration After fixation, tissue should
- Made up of gland, Urgent biopsies & differentiation -Hemolysis be transferred directly to
ethyl alcohol, -Preserves nucleoprotein and -Polarization except at -70’C alcohol
glacial nucleic acid -Harden tissues excessively
acetic acid and -Preserves cytoplasmic & distorts tissue
chloroform granules -Small tissue morphology -Shrinkage
fragments,
curreting and biopsy
materials -Shortened
processing time
GENDRE’S 24hrs -Fast fixation -Gross hardening of
FIXATIVE -Better preservation of tissues -Partial lysis of
(ALCOHOLIC glycogen -Useful for sputum RBC
FORMALIN) (coagulates mucu) -Poor preservation of iron
containing pigments
NEWCOMER’S Made up of -Recommended for
FLUID isopropyl alcohol Mucopolysaccharide and nuclear
protein
-Produces better reaction in
faulgen stain than
Carnoy’s
-Nuclear & histochemical fixative
 OTHER FIXATIVE
FIXATIVE NOTE USE FIXATION ADVANTAGE/S DISADVANTAGES PRECAUTION/S

TIME
LILLIE’S FIXATIVE -LEAD -Used for preservation of 12 – 24hrs -Fixes Connective tissue -Forms insoluble lead
FIXATIVE Glycogen, and Mucin carbonate
Mucopolysaccharide and
Amyloid
GLACIAL ACETIC -GLACIAL 24hrs @ -Fixes precipitates -If combined with
ACID ACETIC ACID 17’C nucleoproteins, chromosome potassium dichromate, lipid
FIXATIVE & chromatin fixing
property is destroyed
ACETONE -Used in fixing brain tissues - Dissolves fat - Preserves glycogen poorly
for the diagnosis of rabies - Evaporates rapidly
OSMIUM -OSMIC ACID -Used extensively for -Electron Microscopy -Expensive
TETROXIDE FIXATIVE neurological tissues -Preserves cytoplasmic -Poor penetration
structures well -Form black deposit due to
-Fixes fats reduced sunlight
-Inhibit hemolysis -Extremely volatile
FLEMMING’S -Most common -Fixes fat
SOLUTION chrome-osmium -Less amount of solution
acetic acid is reuired
fixative

FORMULA
Chromic acid
Glacial acetic
acid Osmium
tetroxide
FLEMMING’S -Recommended
SOLUTION W/O for cytoplasmic
ACETIC ACID structures

FORMULA
Chromic acid
Osmium tetroxide
HEAT FIXATION -Involves -For frozen tissue sections and -Preserves nuclear & -Produces tissue shrinkages &
thermal preparation of bacteriologic cytoplasmic details distortion
coagulation of smear
tissue proteins
TRICHLOROAC -Sometimes
ETIC ACID incorporated into

compound
fixatives
CHAMPY’S FLUID -Recommended
for mitochondria,

golgi, fats

FORMULA
Osmium
tetroxide
Chromic acid
Potassium
dichromate