NAME: NATHALIE A.

SEPE SUBJECT: GENPATH 1

COURSE & BLOCK: BS MEDTECH 3-C DATE: 01-04-2024

CHAPTER 17: COMMON STAINING SOLUTIONS

STAINS PURPOSE/USES

HEMATOXYLIN  most commonly used for routine histologic studies

 recommended for progressive staining of tissues, (i.e. staining for a predetermined time
Aluminum Hematoxylin to adequately stain the nuclei but leave the background tissue relatively unstained, to be
Solutions later counterstained with eosin, Congo red or safranin)

 used for regressive staining and differentiated with I % hydrochloric acid in 70%
Ehrlich’s Hematoxylin alcohol (acid alcohol) until the nucleus is selectively stained

 good regressive stain that may either be used immediately or stored for future use, since
it remains stable for a long time (about 6 months).
Harris Hematoxylin  used for routine nuclear staining, in exfoliative cytology, and for staining of sex
chromosomes

Cole’s Hematoxylin  recommended for routine purposes, especially used in

sequence with Celestine blue

 used as a nuclear counterstain to demonstrate the presence of cytoplasmic glycogen by

special stain
Mayer’s Hematoxylin  used in instances when acid-alcohol differentiation might destroy or decolorize the
stained cytoplasmic
 used in Celestine Blue hemalum method of nuclear staining

Iron Hematoxylin Solutions  used only for differential or regressive staining, using acid-alcohol as a differentiating
agent

Regaud’s Hematoxylin for  most permanent and the simplest is Regaud's modification of iron hematoxylin on
Mitochondria sections of material fixed in potassium dichromate and formalin and subsequently
mordanted in dichromate

Weigert’s Hematoxylin  used in the laboratory, especially for demonstrating muscle fibers and connective tissues
Solution

Heidenhain’s Hematoxylin  a popular cytological stain, especially for the study of mitosis.
 a cytological stain recommended for regressive staining of thin sections

Phosphotongstic Acid  usually demonstrates structures in paraffin as well as celloidin and frozen sections
Hematoxylin (PTAH)

EOSIN  the most valuable stains used for differentially staining connective tissues and
cytoplasm
  most commonly used fixative
Yellowish (Eosin Y)  used as a 1% solution for 15 seconds to 3 minutes, depending on the tissue, type of
fixative and intensity of color desired

 used for human-medical cell diagnosis and serves the histological and clinico-
5 % Aqueous Eosin Y cytological investigation of sample material of human origin

Eosin, Stock Alcoholic Solution  demonstrates various tissue components more dramatically

Eosin-Phloxine B Solution  used for primary staining as a counterstain to Dako Gill's 3 Hematoxylin

 used to examine blood or bone marrow samples

Romanowsky Stains  suited for the examination of blood to detect bloodborne parasites like malaria

CHAPTERS 18: STAINING OF CARBOHYDRATES

STAINS RESULTS

10% Neutral Buffered Formalin or  PAS-positive substances red or magenta red

Bouin’s Solution  Nuclei blue

Helly’s Fluid  Nuclei blue

 Glycogen red

 Nuclei blue or grayish blue

Best Carmine Stain  Glycogen pink to red
 Mucin weak red

Metachromatic Toluidine Blue Stain  Mucopolysaccharide red purple

 Tissue background blue

Alcian Blue Stain  Acid mucins blue

 Nuclei red

 Acid mucins blue

Alcian Blue-PAS Stain  Neutral mucins magenta
 Nuclei pale blue

Gomori’s Aldehyde Fuchsin Stain  Sulfate mucins purple

 carboxylate forms blue

Cryptococcus Neoformans  Mucins red

(Mucicarmine Stain)  Nuclei blue
 Background unstained

Fluorescent Acridine Orange  Acid mucopolysaccharides Black

 Fungi Greenish red fluorescence
 Background Reddish orange fluorescence
 CHAPTER 19: STAINING OF LIPIDS

STAINS RESULTS

Hematoxylin & Eosin Stain  Adipose tissue or fat white and brown
 Lipochrome (lipofuscin) brown pigments

Sudan Black  Lipids blue black

 Nuclei Red

Sudan IV (Scharlach R) Stain  Lipids (mainly triglycerides) – red

 Nuclei – blue/black

Oil Red O  Lipid red

 Nuclei blue

Osmic Acid Stain  Nuclei yellow-orange

 Fats black

CHAPTER 20: STAINING OF PROTEINS AND NUCLEIC ACIDS

STAINS RESULTS

Hematoxylin & Eosin Stain (H&E)  Most of the cytoplasm is eosinophilic

 Red blood cells are stained intensely red

Peracetic Acid-Alcian Blue  cystine and cysteine is stained blue-green

Feulgen Stain  DNA red purple

 Cytoplasm green

 DNA (Chromatin) Green or blue-green

Methyl Green-Pyronin Stain  RNA (Nucleoli) Rose-red
 Granules Dark rose-red
 Plasma cell cytoplasm Purple

Fluorescein  Blue in color

Rhodamine  orange-red emission

Acridine Orange  DNA emits a yellow-green fluorescence

 RNA is stained brick to orange-red

Acriflavine  DNA is stained by a fluorescent yellow color

CHAPTER 24: STAINING OF BONE MARROW AND BLOOD ELEMENTS

 STAINS RESULTS

 Erythrocytes yellowish-red
 Polymorphonuclears: Nucleus dark purple
 Polymorphonuclears: Granules reddish-lilac
 Polymorphonuclears: Cytoplasm pale-pink
 Eosinophils: Nuclei blue
Wright’s Stain  Eosinophils: Granules red to orange-red
 Eosinophils: Cytoplasm blue
 Basophils: Nucleus purple to dark blue
 Basophiles: Granules very dark purple
 Lymphocytes: Nuclei dark purple
 Lymphocytes: Cytoplasm sky blue
 Platelets violet to purple granules

 Bile pigments green

 Collagen, muscle, bone pale pink
 Micro-organisms fungi, parasites purplish-blue
 Starch granules, cellulose sky blue
Giemsa Stain  Pigments (native color is yellow/brown, or green if fixed in dichromate
 containing fixative
 Nuclei dark blue to violet
 Erythrocytes salmon pink
 Cytoplasm varying light blue shades

Wright-Giemsa or Jenner-Giemsa  Nuclei purple/blue

Stain  Cytoplasm pink/blue
 Eosinophils pink/red

 Myeloid cells (except basophils) are positive (i.e., peroxidase activity is shown by
the presence of green to dark blue granules in the cytoplasm).
 Eosinophils are stained most intensely and are often tinged brown-black or green-
black.
Myeloperoxidase (MPO) Stain
 The cytoplasm of neutrophils is filled with blue dye.
 Basophils, lymphocytes and erythroblasts are negative. Monocytes show slight
peroxidase activity.

 Osteoid seams red

Masson’s Trichrome Stain  Mineralized bone blue
 Nuclei dark gray

CHAPTER 25: STAINING OF CONNECTIVE TISSUE

STAINS RESULTS

Gomori’s Silver Impregnation Stain  Reticulin fibers black

 Reticulin fibers Black

Reticulin Stain-Gordon and Sweets’  Nuclei Black
Stain  Background Red
  Nuclei brownish black to black
Van Gieson’s Stain  Collagen (fibrous connective tissue) pink or deep red
 Muscle, Cytoplasm, RBC and Fibrin Yellow

 Muscle, RBC and keratin red

Masson’s Trichrome Stain  Nuclei blue-black
 Collagen and mucus blue

Gomori’s One-Step Trichrome Stain  Muscle fibers red

 Collagen green
 Nuclei blue to green

 Nuclei and elastic fibers Black

Russell’s Modification of the Movat  Collagen Yellow
Pentachrome Stain  Ground substance and mucin Blue
 Fibrinoid, fibrin Intense red
 Muscle Red

 Nuclei, fibrin, muscle fibers red

 Collagen blue
Mallory’s Aniline Blue Stain  Cartilage, bone, mucus varying shades of blue
 Blood, myelin yellow
 Elastic fibers pale pink/yellow or unstained

 Amyloid connective tissues and mucous colloid deep blue stain

Azocarmine Stain  Nuclei red

 Elastic tissue yellowish fluorescence in ultraviolet light (blue if unstained)

Elastic Stain  In the presence of ferric salts (oxidizers), elastic fibers stain with basic fuchsin,
with or without resorcin.

 Elastic fibers appear brown to purple or blue-black with methyl violet on a clear
Weigert's Resorcin-Fuchsin Elastic background. Other structures are colored depending on the counterstain used.
Tissue Stain  The nuclei may be stained red with carmine before or after staining of fibers.

 Nuclei brown - black

Van Gieson Stain  Collagen red
 Muscle, RBCs, cytoplasm yellow

 Elastic fibers black

Verhoeff’s Elastic Stain  Nuclei gray to black
 Collagen red
 Cytoplasm and muscle yellow

 Elastic fibers blue-black to black

Verhoeff-Van Gieson (VVG)  Nuclei blue to black
 Collagen red
 Other tissue elements - yellow

Aldehyde Fuchsin Elastic Stain  Elastic fibers deep blue to purple

 Other tissue elements green
  Elastic fibers purple
Luna Stain  Mast cells purple
 Nuclei black
 Background yellow

Orcein (Taenzer-Unna Orcein Stain)  Elastic fibers dark-brown

 Nuclei blue

 Elastic fibers bright red

Krajian’s Stain  Fibrin and connective tissues dark blue
 RBC orange-yellow

 Basement membranes black

Jone’s Silver Methanamine  Nuclei blue
 Background pink

 Nuclei blue
 Erythrocytes yellow
MSB Stain  Muscle red
 Collagen blue
 Fibrin red (early fibrin may stain yellow, and very old fibrin may stain blue)

 Fibrin, muscle striations, neuroglia and amoebae - dark blue

 Nuclei, cilia, red blood cells blue
Phosphotungstic Acid Hematoxylin  Myelin lighter blue
(PTAH)  Collagen, osteoid cartilage, elastic fibers deep brownish red
 Cytoplasm pale pinkish brown

 Amyloid bright "apple-green" birefringence

Congo Red  Sections that are too thin faint red colors
 Too thick sections yellow birefringent colors

Alkaline Congo Red  Amyloid, elastic fibers, eosinophil granules red

 Nuclei blue

High pH Congo Red  Amyloid, elastic tissue, eosinophilic granules red

 Nuclei blue

Crystal Violet  Amyloid - Purplish red

 Nuclei, cytoplasm, connective tissue - shades of violet

Induced Fluorescent Stain with
Thioflavine-T
 Using UV light source (mercury vapor lamp), UGI Exciter filter, BG38 red
suppression filter and K430 barrier filter, amyloid, elastic tissue etc. will exhibit
silver-blue fluorescence.
 Using blue light fluorescence quartz-iodine or mercury vapor lamp with BG 12
exciter filter and K530 barrier filter, amyloid and elastic tissue will exhibit a yellow
fluorescence.

Lieb’s Crystal Violet  Amyloid purplish violet

 Other tissue elements blue

CHAPTER 26: STAINIG OF MUSCLE AND BONE

 STAINS
Mallory’s Trichrome Stain
Gomori’s Trichrome Stain
Gomori Trichrome Stain for Frozen
Muscle
Mallory's Phosphotungstic Acid
Hematoxylin (PTAH)
Periodic Acid Schiff (PAS) Stain
Sudan Black
Oil Red O Stain (IHC World)
Heidenhain's Iron Hematoxylin
Adenosine Triphosphatase (ATPase)
Succinate Dehydrogenase Stain
Nicotinamide Adenine Dinucleotide
Tetrazolium Reductase (NADH)
Stain
RESULTS
 Erythrocytes orange
 Muscle red
 Collagen blue
 Muscle fibers red
 Collagen green
 Nuclei blue to black
 Myofibrils green
 Intermyofibrillar material bright red
 Nemaline rods, ragged red fibers red on a blue/green background
 Muscle, neuroglia blue
 Nuclei deep blue
 Cytoplasm pale pinkish brown
 Fibrin blue
 Collagen deep brownish red
 Coarse elastic fibers bluish
 Myelin blue to blue-gray
 Type 2 myofibers stain darker with this stain than type 1 fibers
 Intracellular lipid blue-black
 Type 1 myofibers stain darker than the others
 Lipids red
 Nuclei pale blue
 Muscle striations, mitochondria, myelin, and chromatin are stained grey black with
Heidenhain's iron hematoxylin.
 pH 9.4 ATPase: Type 1 fibers light, Type2 fibers dark, Type 2C fibers intermediate
 pH 4.6 ATPase: Type 1 fibers darkest, Type 2B and C intermediate, Type 2A
lightest
 pH 4.3 ATPase: Type 1 fibers darkest, Type 2C fibers intermediate, Type 2A and 2B
fibers lightest
 Oxidative fibers have a relatively dense, purple speckled appearance.
 Nonoxidative fibers have only scattered purple speckles.
 Ragged red fibers, features of mitochondrial myopathies, typically show a dense
rim of DH stain positivity.
 Type I myofibers are darker than the type II myofibers

Myophosphosporylase Stain
Schmorl's Picro-Thionin Stain
Alizarin Red S
Von Kossa Stain
CHAPTER 27: STAINING OF NERVOUS TISSUE
STAINS
Cresyl Fast Violet (Nissl) Stain
Cajal's Gold Sublimate Stain
Modified Holzer's Stain
Weigert-Pal Stain
Luxol Fast Blue-H & E Stain
Luxol Fast Blue-PAS-Hematoxylin
Stain
Weil’s Stain
Baker’s Chromate-Acid Hematin
Stain
 A negative reaction is yellow while polysaccharide of 8 to 12 units gives a reddish
color, followed by various transitional colors as the length of the chain increases.
 Chain lengths of 30 to 35 units give a blue color.
 With myophosphorylase reaction, Type II fibers, particularly
 type 2b, are dark while type I fibers are pale.
 Lacunae and canaliculi dark brown-black
 Bone matrix yellow or brownish-yellow
 Cells red
 Calcium deposits (except oxalate) orange-red. This precipitate is birefringent.
 Calcium salts black or brown-black
 Nuclei red
 Cytoplasm pink
RESULTS
 Nissl substance purple-dark blue
 Neurons pale purple blue
 Cell nuclei purple blue
 Astrocytes bluish black on a light brownish background
 Nerve Cells red
 Nerve fibers unstained
 Glial fibrils blue
 Nuclei pale blue
 Background colorless
 Myelin Sheath blue black
 Cells brown
 Myelin blue-green
 Nuclei dark blue
 Cytoplasm various shades of pink
 Myelin blue
 Fungi and PAS-positive elements rose to red
 Nuclei dark blue
 Cytoplasmic nucleoproteins bluish purple
 Capillaries red
 Myelin black
 Background yellow
 Myelin Sheaths, Mitochondria, and Erythrocytes dark blue
Swank & Davenport’s Marchi Stain  Degenerating Myelin (Early and Late Products) Black
 Background: colorless to pale brown

 Axons Brown to black

Microwave Modification of  Cytoplasmic neurofibrils Brown to black
Bielschowsky’s Stain  Neurofibrillary tangles and Plaques Dark brown or black
 Neuromelanin Black
 Lipofuscin Brown or black

 Large and small peripheral neurites black

 Axons black
Sevier-Munger Stain  Myelin sheath light brown
 Neuritic plaques and tangles black
 Argentaffin granules black

 Myelin blue
Methylene blue-azure II-basic  Other tissue elements light blue
Fuchsin  Collagen pink/red
 Elastin red

CHAPTER 28: STAINING OF MICROORGANISMS

STAINS RESULTS

Gram Stain  Gram positive organism blue-black

 Gram negative organism red

 Gram positive organisms, fibrin, some fungi blue

Modified Brown-Brenn Stain  Gram-negative organism red
 Nuclei red
 Other tissue elements yellow

 Gram positive organism blue-black

 Gram negative organism pink-red
Gram-Twort Stain  Nuclei red
 RBCs and most cytoplasmic structures green Elastic fibers black

Acid-Fast Stain  acid-fast cells will be reddish-pink

 non-acid fast cells will be blue

Ziehl-Neelsen Stain  Mycobacteria, some fungal organisms red

 Background blue

Fite Stain  M. leprae and other acid-fast bacteria bright red

 Background light blue

Microwave Auramine-Rhodamine  Acid-fast organism Reddish-yellow fluorescence

Fluorescent Stain  Background Black

Toluidine Blue Stain  Helicobacter- dark blue against a variably blue background
 Cresyl Violet Acetate Stain
Dieterle Stain
Warthin-Starry Stain
Steiner and Steiner Microwave
Stain
Grocott Methenamine Silver Nitrate
(GMS) Stain
Lendrum's Phloxine-Tartrazine
Stain
Orcein Stain
Giemsa Stain
Rapid Giemsa Stain
 Helicobacter and nuclei blue violet
 Background shades of blue-violet
 L. pneumophilia, spirochetes, bacteria Brown to black
 Background Pale yellow or tan
 Spirochete black
 Background golden yellow
 Spirochetes Dark brown to black
 H. Pylori Dark brown to black
 L. Pneumophilia Dark brown to black
 Other non-filamentous bacteria Dark brown to black
 Background light yellow
 Fungi- Cell walls should be crisp black, and the internal structures should be
visible.
 Mucin Taupe to dark gray
 Background Green
 Viral inclusions bright red
 Red blood cells variably yellow to orange red
 Nuclei blue-gray
 Background yellow to pink
 Hepatitis-B antigen, elastic, some mucins brown-black
 Background yellow
 Protozoa and some other organism dark blue
 Background pink-pale blue
 Nuclei blue
 Bacteria blue
 Mast cell granules deep blue
 Eosinophilic granules red
 Nuclei blue
 Cytoplasm pink