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PHT LAB T 11-2


Formaldehyde Usually buffered to 10% formalin is a Formalin (Most widely 24 hours
pH7 with phosphate 1:10 dilution of used fixative)
buffer 100% formalin in
10% Formol A simple Formaldehyde Recommended for  24 hours
Saline microanatomical 40% = 100mL  fixation of central at 35C(95F)
fixative made up of NaCl = 9gm  nervous tissues and  48 hours
saturated Distilled water = general post mortem at 20-25C
formaldehyde (40% 900mL tissues for (65-77F)
by weight volume) histochemical
diluted 10% with examination.
sodium chloride.
10% Neutral recommended for  Sodium best fixative for : 4 - 24
Buffered preservation and dihydrogen tissues containing hours
Formalin or storage of surgical, phosphate iron pigments and
Phosphate- postmortem and (anhydrous) 3.5 elastic fibers which
Buffered (pH 7) research specimen gm  Disodium do not stain well after
hydrogen Susa,
(anhydrous) 6.5
gm 
100 mL 
Distilled water
900 mL
Formol-Corrosive penetrates small  Sat. Aq. Formol-mercuric : 3 - 24
(Formal- pieces of tissues Mercuric chloride solution is hours
Sublimate) rapidly - no need for chloride 90 mL  recommended for
"washing- out" --- Formaldehyde 10 routine post- mortem
can be transferred mL tissues
directly from fixative
to alcohol
Alcoholic Formali can be used for  95% ethyl alc. good for preservation Post-
(Gendre’s) rapid diagnosis --- it saturated w/ of glycogen and for fixation
FixativE fixes and picric acid 80 mL microincineration with
dehydrates at the  strong technique phenol-
same time formaldehyde formalin for
soln. 15 mL  6 hours or
glacial acetic more
acid 5 mL
Glutaraldehyde • made up of 2 2.5% soln. - used used for electron : 1/2 - 2
formaldehyde for small tissue microscopy --- hours
residues, linked by fragments and buffered
three carbon chains needle biopsies glutaraldehyde then
fixed in 2-4 hours osmium tetroxide
@ room temp. •
4% soln. -
for larger tissues
less than 4 mm
thick fixed in 6-8
hours up to 24
Mercuric Most common Mercuric Widely used as a
Chloride metallic fixative --- chloride 5 gm secondary fixative
frequently used in Potassium
saturated aqueous dichromate 2.5
solutions of 5%-7% gm Sodium
sulfate (optional)
1 gm Distilled
water 100 mL
Zenker’s Fluid Mercuric deposits Made up of Good general fixative : 12-24 hrs
may be removed by mercuric for adequate
immersing tissues chloride stock preservation of all
in ALCOHOLIC solution plus kinds of tissues and
IODINE prior to GLACIAL give excellent
staining, through a ACETIC ACID --- staining results
process known as added before its
DEZENKERIZATION. use to prevent
turbidity and
formation of dark
Zenker-Formol Brown pigments are Stock solution, An excellent 12-24 hrs
(Kelly’s Solution) produced if tissues Mercuric microanatomic
are allowed to stay chloride 5gm fixative for pituitary
in the fixative for Strong gland, bone marrow,
>24 hrs. due to RBC formaldehyde, and blood containing
lysis 40% 5 mL organs --- spleen and
Heidenhain’s An excellent Mercuric Recommended 3-12 hrs.
Susa Solution cytologic fixative chloride 45 gm mainly for tumor
Sodium chloride biopsies especially
5 gm the skin.
acid 20 gm
Glacial acetic
acid 40 mL
40% 200 mL
Distilled water
800 mL
B-5 Fixative Two working Distilled water 90 Commonly used for Rapid
solutions are kept cc Mercuric bone marrow fixation can
separate, since the chloride 6 gm biopsies. be achieved
mixture is unstable. Sodium acetate in 1 1/2 – 2
1.25 gm (Just hrs
prior to use, add
1 cc of
40% for 10 cc of
B-5 fixative.)
Chromic Acid A strong oxidizing Used in 1% - 2% Precipitates all
agent --- in need of aqueous solution proteins and
strong reducing --- usually a adequately preserves
agent before use in constituent of a carbohydrates.
order to prevent compound
counteracting fixative.
Potassium Fixes but does not Used in 3% Preserves lipids
Dichromate preserve aqueous solution
Regard’s Potassium Recommended for 12-48 hrs.
(Muller’s) Fluid dichromate 80 demonstration of
mL Strong chromatin,
formaldehyde, mitochondria, mitotic
40% 20 mL (To figures, golgi bodies,
be added just RBC, and
before use) colloidcontaining
Orth’s Fluid Recommended for Potassium di Preserves myelin : 36-72 hrs.
study of early chromate 2.5 % better and
degenerative 100 mL Sodium demonstrates
process and tissue sulfate (optional) rickettsiae
necrosis. 1 gm Strong
40% 10 mL
Lead Fixatives Takes up CO2 to Used in 4% Recommended for
form insoluble lead aqueous solution acid
carbonate on of basic lead mucopolysaccharides
prolong standing --- acetate.
removed by: a)
Filtration b)
Addition of acetic
acid --- drop by drop
to lower the pH and
dissolve the
Bouin’s Solution Tissue fixed with Sat. solution of Recommended for 6-24 hrs
this fixative retain picric acid 75 mL fixation of embryos
little affinity for Strong and pituitary biopsies
basic dyes. formaldehyde
40% 25 mL
Glacial acetic
acid 5 mL
Brasil’s Alcoholic Formaldehyde An excellent fixative Overnight
Picroformol 37% 2040 mL for glycogen tissue
Fixative Picric acid 80 gm fixation by
Ethanol or automatic
Isopropyl alcohol processing
6000 mL technique
Trichloroacetic may utilize
acid 65 gm 3-4 changes
of Brasil’s
fixative at ½
to 2 hours
Glacial Acetic Acetic acid is used in Solidifies at
Acid conjunction with 17oC ---
other fixatives to form Glacial
a compound solution. acetic acid
Methyl Alcohol Fixes and • Excellent for fixing If left in
100% dehydrates at the dry and wet smears, fixative for
same time blood smears and more than
bone marrow tissues 48 hrs. ---
tissues may
and difficult
to cut.
Isopropyl • Used for fixing
Alcohol 95% touch preparations
Ethyl Alcohol • Lower Used at conc. of Used as a simple 18-24 hrs.
concentrations --- 70% - 100% fixative
RBC’s become
hemolyzed and
WBC’s are
Carnoy’s Fluid Rapid in action Absolute alcohol Recommended for : 1-3 hrs
60 mL fixing chromosomes,
Chloroform 30 lymph glands and
mL Glacial acetic urgent biopsies.
acid 10 mL
Newcomer’s Acts both as Isopropyl alcohol Recommended for 12-18 hrs.
Fluid anuclear and 60 mL Propionic fixing at 3C
histochemical acid 30 mL mucopolysaccarides
fixative. Petroleum ether and nuclear proteins.
10 mL Acetone
10 mL Dioxane
10 mL
Osmium A poor penetrating A pale yellow Fixes conjugated-fats
Tetraoxide agent and very powder and lipids
(Osmic Acid) expensive dissolves in permanently.
water to form a
strong oxidizing
Flemming’s Most common Aqueous Recommended for 24-48 hrs
Solution chrome-osmium chromic acid 1% nuclear preparation
acetic acid fixative 15 mL Aqueous of each sections.
used. osmium
tetroxide 2% 4
mL Glacial acetic
acid 1 mL
Flemming’s Made up only of Aqueous Recommended for 24-48 hrs
Solution without chromic and some chromic acid 1% cytoplasmic
acetic acid osmic acid 15 mL Aqueous structures
osmium particularly the
tetroxide 2% 4 mitochondria
Tricholoroacetic Poor penetrating Incorporated into Precipitates proteins
Acid agent compound and may be used as a
fixatives weak decalcifying
Acetone Recommended for Used in fixing brain Used at ice
the study of water tissues for diagnosis cold
diffusible enzymes - of rabies. temperature
-- phosphatases and ranging
lipases from -5 to 4