LEC#13 HISTOPATHOLOGY

PROF. Dr. Marissa A. Cauilan

LEC MLS – 3A
S.Y. 2022-2023
STAINING

PRINCIPLES OF STAINING: Xylene

STAINING  Is not miscible with aqueous solutions and low graded

alcohol
●Process whereby tissue components are made visible in
microscopic sections by direct interaction with a dye or staining  Should therefore be subsequently removed with
solution absolute alcohol, followed by descending grades of
alcohol to prevent damage and detachment of sections.
●Enhance contrast and visualization of cell or its components
1.Deparaffinization
●Colored compound is used to produce contrast based on
varying affinities of cell components -Immersing the paraffin section in xylene 2x
○ colored produced by the tissues or structure are not -After cutting, tissue will be embedded in a paraffin block so it's
the actual color of the tissue itself. The color produced important that we now remove the paraffin because paraffin is
is based on the AFFINITY of that structure to the dye. not visible with the stain since stain is usually in aquarius or
alcoholic solution.
○ Basic dye has strong affinity for acidic structure like
nucleus, hence, basic dyes stain the nucleus blue. In -We have to remove the paraffin using xylene. It will be
contrast, the acidic dye has greater affinity for basic immersed in xylene two times followed by rehydration.
structures for example the cytoplasm. acidic dye will
stain the cytoplasm since cytoplasm has a greater 2.Rehydration
affinity for acidic dye.
-Absolute alcohol followed by descending grades of alcohol
○ Basic dye → acid structure
- paraffin will be removed with xylene then remove the
○ Acid dye → basic structure xylene using alcohol and replacing it with water

3.Staining

HISTOLOGIC STAIN -Immersing the sample in dye solution

●Purified form of a coloring agent or crude dye that is generally 4.Dehydration

applied in aqueous solution
-Increasing grades of alcohol
●Individual variation in properties of tissue constituents
5.Clearing
produce variation in color
-Two changes of xylene
○ that is why it is important to have a contrast
between the different stains to enable to differentiate Alcohol-based stains: no need to immerse in water, just the
between the structure descending grades of alcohol kaya tinatawag siyang na
MORDANT METHOD OF STAINING:
●Chemical compound that reacts with the stain to form an
insoluble, colored precipitate on the tissue and make staining
reaction possible DIRECT STAINING

○ this mordant act as a bridge because it forms a ●Process of giving color to the sections by using aqueous or
precipitate so it forms a mordant dye “lake” complex alcoholic dye solutions

○ Mordant will also make counterstaining and ●One dye is used

decolorization possible because usually the dye per
se can stain the tissue but once decolorized or ●Basic dyes-positive charge (giving a greater affinity to
counterstain the stain may be easily removed negatively charge structures)
therefore with the use of mordant it will intensify the ●Ex: Methylene blue
stain because it will form an insoluble colored
precipitate. INDIRECT STAINING
STAINING OF PARAFFIN SECTIONS:  Action of the dye is intensified by adding another agent
or a MORDANT which serves as a bridge between the
Paraffin wax tissue and the dye.
 Is poorly permeable to most staining solutions and  Mordant combines with a dye to form a colored “lake”,
shouldtherefore be removed from the section prior to whichin turn combines with the tissue to form a “tissue-
staining. mordant- dye-complex”.

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o This allows subsequent counterstaining and *If the primary dye is a basic dye, the differentiator would be an
dehydration to be carried out easily. acid solution while if the primary dye is an acid dye, the
differentiator would be a basic solution.
o A mordant may be applied to the tissue before
the stain or may include as part of staining
METACHROMATIC STAINING
technique orit may be added to the dye solution
itself.
 It entails the uses of specific dyes which differentiate
o Example: particular substances by staining them with a color
that is different from that of the stain itself.
Potassium alum with hematoxylin in
Ehrlich’s hematoxylin and iron in  Tissues components combine with these dyes to form
Weigert’s hematoxylin a different color from the surrounding tissue.

 It is particularly employed for staining cartilage,

connective tissue, epithelial mucins, mast cell
granules, and amyloids.

 Azure or toluidine blue

 The actual staining process may involve immersing

the sample (before or after fixation and mounting) in
dye solution, followed by rinsing and observation.

METALLIC IMPREGNATION

 Process where specific tissue elements are

PROGRESSIVE STAINING demonstrated,not by stains, but by colorless solutions
of metallic salts which are reduced by the tissue,
 A process whereby tissue elements are stained in a
producing opaque, usually black deposits on the
definitesequence, and the staining solution is applied for
surface of the tissue or bacteria.
specific period of time or until the desired intensity of
coloring of thedifferent tissue elements is attained. o The most valuable metals for the purposes are
gold(gold chloride) and silver (silver nitrate)
o Once the dye is taken up by tissue, it is not
washedor decolorized. VITAL STAINING
 NOTE: The distinction of tissue relies solely on the  Selective staining of living cell constituents,
selectiveaffinity of the dye for different cellular elements.
 Demonstrating cytoplasmic structures by phagocytosis
REGRESSIVE STAINING of the dye particle (cytoplasmic phagocytosis), or by
●Tissue is first overstained staining of pre-existing cellular components (true vital
staining) --- staining of mitochondria by Janus green
●Excess stain is removed or decolorized until desired intensity
of color is attained.  Examples: (i.) vital staining of reticulo-endothelial
system (with Tyrpan blue, or propidium iodide for
●Hematoxylin and Eosin (commonly used) eukaryotic cells)

●Overstained the tissue with hematoxylin then, counterstain it  Purpose: To reveal cytological details (Note: nucleus
with eosin of a living cell is resistant to vital stain that’s why is
NOT demonstrated)
DIFFERENTIAL STAINING
 Note: “Demonstration of nuclear structures during vital
●Selective removal of excess stain from the tissue during staining suggests permeability of the membrane of the
regressive staining dye,signifying the death of the cell.”

●Uses more than one chemical stain to better differentiate INTRAVITAL STAINING
between various microorganisms or structures/cellular
components  Injecting the dye into any part of the animal body
(either intravenous, intraperitoneal or subcutaneous),
●Requires washing or use of acids and oxidizing agents producing specific coloration of certain cells,
particularly those of the reticulo-endothelial system.
●Basic dye-acid solution
 Common dyes used:
●Alcohol acts as a differentiator for both basic and acidic dyes
o lithium, carmine and India ink
●Mordant can act as a differentiating agent

●The differentiating solution is important to produce different

contrast of colors on the tissue that is why it was called
differential staining.

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SUPRAVITAL STAINING ●Cellulose nitrate (celloidin) is soluble in absolute alcohol and

will be removed if absolute alcohol is used in the final
 Used in microscopy to examine living cells that have dehydration prior to clearing of stained sections.
been removed from an organism.

 Supravital stains: those that enter and stain living cells

(examples: New Methylene Blue and Brilliant Cresyl STAINS AND STAINING SOLUTION
Blue for reticulocyte staining --- toxic to the organism)

 Note: (partly) Due to their toxic interaction inside a

living cell, when supravital stains enter a living cell, NATURAL DYES
they might produce a characteristic pattern of staining
- natural dyes are those obtained from plants and animals,
different from the staining of an already fixed cell
previously utilized for dyeing of wool and cotton. Among the
("reticulocyte" look versus diffuse "polychromasia")
most common natural dyes available are:
 Stains are used in very dilute solutions --- ranging from 1.Hematoxylin
1:5,000 to 1:50,000
● natural dye derived by extraction from the core or
o Note: Many stains may be used in both living
and fixed cells. the heartwood of a Mexican tree known as
"Hematoxylin Campechianum”.
 Common dyes used are:
● most valuable staining reagent
1. Neutral red - probably the best vital dye.
● Powerful cytology stain because it has powerful
2. Janus green - especially recommended for nuclear and chromatin staining capacity.
mitochondria.
● polychrome property
3. Trypan blue - one gram of dye is dissolved in 100 ml.
of sterile distilled water to be used immediately; it is ● Hematin – active coloring agent
dangerous to allow the suspension to stand for more
● most frequently used in combination with alum, iron,
than one hour, because it is likely to become toxic to the
chromium, and copper salts, which act as mordants.
cell.
Note:
4. Nile blue

5. Thionine ● Without the mordants we cannot maximize the stain/dye.

6. Toluidine blue ● We need these mordants so that when you decolorize a

counter stain, the structures will still retain the color of
HEMATOXYLIN AND EOSIN STAINING Hematoxylin dye.

 Corner stone of tissue-based diagnosis 2.Cochineal Dyes

 To stain cell nuclei (and other parts) blue and an eosin ● extracted from the female cochineal bug (Coccus
dyeto stain other structures pink or red Cacti), which is treated with alum to produce the dye,
carmine.
 Binds strongly to acids and consequently binds to
nuclear DNA and stains nuclei blue ● powerful chromatin and nuclear stain for fresh
material and smear preparations (best for cytology).
 Most fixatives can be used except osmic acid
solutions which inhibit hematoxylin ● Best’s Carmine stain (glycogen)

All specimens are initially stained using H&E (reason: H&E ● Picocarmine (neuropathological studies)
can confirm the basic tissue type and help to localize the
lesion. Thus, “lesion” means any area of damage, infection, 3.Orcein
inflammation, tumor, necrosis or otherwise abnormal tissue)
● vegetable dye extracted from certain lichens which
FROZEN SECTION STAINING are normally colorless, but which, when treated with
ammonia and exposed to air, produce blue or violet
●stained as in paraffin sections although the duration of colors.
staining is usually shorter.
● weak acid and soluble in alkali
●produces well-differentiated sections that are semi-permanent.
● mainly used for staining elastic fibers.
COLLODIONIZATION OF SECTIONS

●process of coating the slide with dilute (thin) celloidin

solutions.

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SYNTHETIC DYES Ex:

 Romanowsky dyes
Source: derived from hydro-carbon benzene (C6H6) taken
from coal tar  Giemsa’s stain
 Irishman’s stain
-also known as “Coal Tar Dyes” or “Aniline Dyes” COMMON STAINING SOLUTIONS
Chromophores HEMATOXYLIN
-are substances with definite atomic groupings and are ●most commonly used for routine histologic studies
capable of producing visible colors.
●mordants used to demonstrate nuclear end cytoplasmic
Chromogens structures are alum and iron, forming lakes or colored
complexes
-are simple benzene compounds that contain chromopores
Auxochrome ●basic dye
-an auxiliary radical or substance which imparts to the
●stain acidic (or basophilic) structures purplish blue
compound the property of electrolytic dissociation, thereby
altering the shade of the dye, enabling it to form salts with COMMON STAINING SOLUTIONS
another compound, and ultimately retaining its color.
Note:
 Chromogens are different from dyes in that any color ALUMINUM HEMATOXYLIN SOLUTIONS
that they impart to the tissue is not permanent and can,
therefore, be easily removed. ●progressive and regressive staining

●aluminum salts give a blue lake, and increase the selectivity

1. Acid dyes for nuclei, especially if acid is added or is used as a
Active coloring substance is found in acid component differentiating agent
andinactive base ●Ehrlich’s hematoxylin and Harris hematoxylin solutions
Ex: Picric acid- forms salt with an alkaliPicric acid
-is the only substance so far that can fix, differentiate and ●During staining, alum hematoxylin stained sections are
usually passed on to an alkaline solution (e.g. 1% hydroxide) in
stain tissue all by itself
order to neutralize the acid and free the OH group, to form an
-act as counterstain to basic cytoplasmic stains, to acid
insoluble blue aluminum hematin-tissue-lake. Such procedure
fuchsin in Van Gieson's connective tissue staining, or is known as blueing
to crystal violet for the microscopic study of fungi.
--used as fixative ●Blueing with ammonia, lithium carbonate or Scott’s Tap Water
Substitute has more rapid action than tap water
-used as decalcifying agent
-used as tissue softener ○tap water is enough to produce blueing of the hematoxylin
Acidophilic-refers to basic cell structures
EHRLICH’S HEMATOXYLIN
(collagen, eosinophilic granules of leukocytes,
etc.) that have anaffinity for the acid dye ions. ●hematoxylin 2 gm
2. Basic Dyes
●absolute ethyl alcohol 100 m
active coloring substance is found in a basic
component that combines with the acid radical ●aluminum potassium sulfate 15 gm approximately
(usuallytaken from sulfuric, acetic or hydrochloric
acid). ●glycerin 100 ml
Ex: Methylene blue ●distilled water 100 ml
Methylene blue
●glacial acetic acid 10 ml
-for bacterial staining
-both used as indicator and dye ●regressive staining, and differentiated with 1% hydrochloric
Basophilic acid in 70% alcohol (acid-alcohol)
-refers to acidic cell structures (chromatin, mucus,
●mucopolysaccharide substances such as cartilage and
cartilagematrix etc.) have an affinity for basic dye ions
cement lines of bones are also stained intensely blue
3. Neutral Dyes
-formed by combining aqueous solutions of acid ●tissues subjected to acid decalcification
andbasic dyes,
●not an ideal for frozen sections
-insoluble or barely soluble in water but soluble in alcohol
- Ethyl alcohol or acetic acid-fixed tissues, on the
otherhand, readily take in both basic and acidic dyes.

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HARRIS HEMATOXYLIN -Absolute ethyl alcohol ml

●hematoxylin 1 gm SOLUTION B:

absolute ethyl alcohol 10 m ammonium/potassium alum 20 gm -30% anhydrous ferric chloride 4 ml

distilled water 190 ml
-Concentrated hydrochloric acid 1 ml
mercuric oxide (red) 0.5 gm glacial acetic acid 10 ml
-Distilled water 100 ml
●good regressive stain
HEIDENHAIN’S HEMATOXYLIN
●routine nuclear staining, in exfoliative cytology, and for
staining of sex chromosomes ●regressive staining of thin sections - all components are black
or dark grey - black.
COLE’S HEMATOXYLIN
●nuclear and cytoplasmic inclusions such as chromatin,
●hematoxylin 1.5 gm chromosomes, nucleoli, centrosomes, and mitochondria.

1% Iodine in 95% alcohol 50 ml sat. aq. ammonium alum 700 ●Voluntary muscle striations and myelin
ml distilled water 250 ml
-MORDANT DIFFERENTIATOR: Ferric
●alum hematoxylin solution recommended for routine ammonium sulfate 2.5 gm
purposes
-Distilled water 100 ml
●artificially ripened with an alcoholic iodine solution
-HEMATOXYLIN STAIN: Hematoxylin 1.5 gm
MAYER’S HEMATOXYLIN
-95% ethyl alcohol 10 ml
●hematoxylin 1 gm sodium iodate 0.2 gm potassium alum 50
gm citric acid 1 gm chloral hydrate 50 gm distilled water 1000 -Distilled water 90 ml
ml
PHOSPHOTUNGSTIC ACID HEMATOXYLIN (PTAH)
●chemically ripened with sodium iodate
●Nuclei, fibrin, muscle striations, and myofibrils are colored
●regressive stain and progressive stain blue while collagen, bone and cartilage take an orange-red or
brownish red to deep brick-red stain.
●nuclear counterstain to demonstrate the presence of
cytoplasmic glycogen ●Progressive

●mucopolysaccharides -Hematoxylin 1 gm

●celestine blue hemalum method of nuclear staining -Phosphotungstic acid 20 gmDistilled Water 1000 ml

IRON HEMATOXYLIN SOLUTIONS EOSIN

●differential or regressive staining, using acid-alcohol as a ●most valuable stains used for differentially staining connective
differentiating agent. tissues and cytoplasm

●Weigert's Solution, using ferric ammonium chloride, and ●acid dye/acidophilic/ oxyphilic/ eosinophilic
Heidenhain's solution, using ferric ammonium sulfate (iron
●counterstain to hematoxylin
alum) as mordants
●background stain
●Tissue structures are stained blackish or grayish, according to
the extent of differentiation ●Eosin Y and Eosin B
REGAUD’S HEMATOXYLIN ROMANOWSKY STAIN
●demonstrate mitochondria by light microscopy ●based on a combination of eosinate (chemically reduced
eosin) and methylene
WEIGERT’S HEMATOXYLIN SOLUTION
●Wright's stain, Jenner's stain, Leishman stain and Giemsa
●standard iron hematoxylin stain used for demonstrating
stain
muscle fibers and connective tissues
●blood or bone marrow samples
●recommended when the preceding stains contain acid (e.g.
Van Gieson stain containing picric acid) which decolorizes ●blood-borne parasites like malaria.
nuclei stained with alum hematoxylin
- Used in hematology and microbiology
SOLUTION A:

-Hematoxylin 1 gm

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OTHER STAINS 3.Schiff’s Reagent

4.Mallory’s Fuchsin Stain

Acid Fuchsin-Picric Acid (Van Gieson’s Stain) 5.Aldehyde Fuchsin (Gomori’s Stain)

●mixture of picric acid and acid fuchsin for demonstration of BENZIDINE

connective tissues.
●For staining hemoglobin
Acid Fuchsin (Masson Stain)
CARMALUM (MAYER'S) SOLUTION
●stain collagen, smooth muscle, or mitochondria
●mordanted dye acting as a basic dye and staining acidic
●nuclear and cytoplasmic stain in Mallory's trichrome method substances

●Van Gieson's picro-fuchsin, acid fuchsin imparts its red color CELESTINE BLUE
to collagen fibers
●oxazine dye used as an alternative to iron hematoxylin
ACRIDINE ORANGE nuclear stain

●permits discrimination between dead and living cells ●forms a strong staining lake with iron alum, acting as a
(discrimination between dead and living cells is used in mordant to bind hematoxylin
combination with ethidium bromide)
●resistant to strong acid dyes, and is recommended for routine
●green fluorescence for DNA and a red fluorescence for RNA staining of fixed sections, giving a good nuclear definition when
used in conjunction with alum hematoxylin.
●nucleic acid selective fluorescent cationic dye useful for cell
cycle determination CONGO RED

ACRIDINE RED 3B ●is best known as an indicator

●demonstrate deposits of calcium salts and possible sites of ●used to identify deposits of protein in tissue called amyloid.
phosphatase activities
CRESYL VIOLET
ALCIAN BLUE
●stain nervous tissues
●complex, water-soluble phthalocyanin dye, similar to
chlorophyll, which stains acid mucopolysaccharides by forming ●stains the acidic components of the neuronal cytoplasm
salt linkages with them (specifically Nissl bodies) a violet color

●blue color resistant to various counterstaining procedures CRYSTAL VIOLET

●specific for connective tissue and epithelial mucin ●is a nuclear or chromatin stain used for staining amyloid in
frozen sections and platelets in blood
●often combined with PAS
●Gentian violet - staining solution formed by the mixture of
ALIZARIN RED S crystal violet, methyl violet and dexterin.

● forms an orange-red lake with calcium at a pH of 4.2 ETHIDIUM BROMIDE

ANILINE BLUE ●intercalates and stains DNA, providing a fluorescent red-

orange stain
●cytoplasmic stain used for counterstaining of epithelial
sections. ●marker for apoptosis in cells populations

AZOCARMINE ●locate bands of DNA in gel electrophoresi

●Nuclei are deep red, cytoplasm is pale red ●used in conjunction with acridine orange (AO) in viable cell
counting - live cells to fluoresce green whilst apoptotic cells
BASIC FUCHSIN retain the distinctive red-orange fluorescence.
●deep staining of acid- fast organisms, for mitochondria, for GIEMSA STAIN
differentiation of smooth muscles with the use of picric acid
●mixture of methylene-blue and eosin
●Feulgen's and Schiff's reagent for the detection of aldehydes
●staining blood to differentiate leukocytes
●Van Gieson's solution for connective tissues, mucin, and for
elastic tissue staining ●mostly used on methanol-fixed blood films

1.Carbol-Fuchsin ●It also binds to some pathogens, including spirochetes

(syphilis), trypanosomes (sleeping,
2.Coleman’s Feulgen Reagent

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sickness and Chagas disease) and plasmodium(malarial homologues of the dye (azures) and deaminized oxidation
parasite). products (thiazoles).

GOLD SUBLIMATE ● methylene blue - methylene blue, azures and thiazoles.

●stain used for metallic impregnation, made up of gold chloride ● stains nuclei blue while cartilage matrix, mucin, mast cell
and mercuric chloride. granules and connective tissues generally take a reddish-violet
color.
IODINE
●Plasma cells
●stains amyloid, cellulose, starch, carotenes and glycogen
●cytological examinations of fresh sputum for malignant
●widely used for removal of mercuric fixative artefact pigments, cells
and as a reagent to alter crystal and methyl violet so that they
may be retained by certain bacteria and fungi. ●bacterial

●aqueous or alcoholic solutions ●diagnosis of diphtheria

GRAM’S IODINE ●vital staining of the nervous tissue NILE RED

●used to identify and differentiate bacteria ●formed by boiling Nile blue with sulfuric acid

● Gram-positive - staphylococci, streptococci and ● lipophilic stain; it will accumulate in lipid globules
pneumococci gram-positive inside cells, staining them red

●Gram-negative - coliforms and Neisseria OIL RED O

LUGOL’S IODINE ●multiple sclerosis - macrophages take up the lipid-rich

debris and stain strongly with this dye
● brown solution that turns black in the presence of
starches ● identify neutral lipids and fatty acids in smears and
tissues
●mordant in Gram's staining
●Fresh smears or cryostat sections of tissue
JANUS GREEN
●identifying fat emboli in lung tissue or clot sections of
● used for demonstrating mitochondria during intravital peripheral blood
staining
ORCEIN
MALACHITE GREEN
●excellent stain for elastic fibers
●weakly basic dye used as a contrast stain for staining ascaris
eggs and erythrocytes, and as a bacterial spore stain ●dermatological studies - ability to demonstrate the finest
and most delicate fibers in the skin
●used both as a decolorizer and as a counterstain
OSMIUM TETROXIDE
●primarily a counterstain
●selective stain for unsaturated lipids and for
MASSON’S TRICHROME lipoproteins such as myelin
●three-color staining protocol ●Fat, which reduces osmium tetroxide to osmium dioxide, is
stained black, and may be demonstrated from the
●distinguish cells from surrounding connective tissue
tissue by using chrome-osmium solutions or by the frozen
●produce red keratin and muscle fibers, blue or green staining section method
of collagen and bone, light red or pink staining of cytoplasm,
PERIODIC ACID SCHIFF (PAS)
and black cell nuclei.
●stains glycogen, mucin, mucoprotein, glycoprotein, basement
METHYL GREEN
membranes, capsules, and blood vessels as well as fungi and
●stains chromatin green in the presence of an acid intracellular carbohydrates such as glycogen in hepatocytes

●false positive reactions with mucin ●Cells that secrete mucus

METHYLENE BLUE PHOSPHOTUNGSTIC ACID

●contains some azures or methylene violet ●common negative stain for viruses, nerves,
polysaccharides, and other biological tissue materials.
● stains acidic cell parts (like the nucleus) blue and is a good
counterstain with Eosin Y ●demonstration of striated muscle fibers and
mitochondria, which stain blue.
●"Polychroming" involves the oxidation of methylene blue,
resulting in loss of methyl groups and leaving lower
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PICRIC ACID ●Azurophilia (affinity for the oxidation products of methylene

blue called azures, which are reddish purple)
●contrast stain to acid fuchsin, for the demonstration of
connective tissue (Van Gieson's stain) ●Acidophilia ( affinity for eosin)

●cytoplasmic stain in contrast to basic dyes ●neutrophilia (affinity for a complex of dyes in the mixture,
which are pale lilac)
●counterstain to crystal violet
●Usually for Hematology
●tissue fixative and decalcifying agent.

PRUSSIAN BLUE
OIL SOLUBLE DYES (LYSOCHROMES)
● is an insoluble colored salt of ferric ferrocyanide (an
iron cyanide compound)

●microanatomical color contrast of specimens Sudan Black

●demonstration of the blood and lymph vessels by injection ●Most sensitive of the oil soluble dyes
(intravital staining)
●greater affinity for phospholipids than other lysochromes -
RHODAMINE B coloring neutral lipids (triglycerides) by simple dissolution of the
dye.
●used with osmic acid to fix and stain blood and glandular
tissues ●ability of fats to absorb Sudan Black is related to dye
concentration, temperature and physical state of the fats.
SAFRANIN
Sudan IV
●produces red nuclei
●has no secondary amino group
●used primarily as a counterstain
●does not color phospholipids or the fine lipid droplets
●give a yellow color to collagen
●recommended for staining triglycerides ( neutral lipids), giving
Silver Nitrate them a deep and intense red stain
●used in 10% aqueous solution to prepare various dilutions to Sudan III
be used in identification of spirochetes, reticulum and other
fiber stains. ●good as a fat stain for central nervous system tissues,

●In Histopath it is more on the reticulum, which it stains black. ●giving a less deep and lighter orange stain compared to the
darker staining Sudan IV.
Toluidine Blue
CHIEF SOLVENTS USED FOR STAINS
●It is recommended for staining of Nissl granules or
chromophilic bodies. 1.WATER - most common

●stains nucleic blue, and can be used to differentiate different 2.ALCOHOL - most common
types of granules ( e.g. within mast cells).
3.ANILINE WATER
●preliminary stain to identify sections that will later be
examined by electron microscopy. 4.PHENOL

Van Gieson Stain

●binds to collagen in the extracellular matrix, staining it pink.

●Often it is combined with a stain for elastic fibers (elastic van

Gieson) which stain black, allowing the two major elements of
connective tissue to be differentiated.

Victoria Blue

●used for demonstration of neuroglia in frozen sections.

Von Kossa Stain

●silver reduction method that demonstrates phosphates and

carbonates.

Wright Stain

●Basophilia (affinity for methylene blue)

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