THEORY OF

STAINING

By Group B
PURPOSE OF STAINING
 To see organism better
 To differentiate one organism from another
 To determine particular structures
 TYPES OF STAINING
PROCESS
1. Vital staining: staining of structure in living cells, divided into:
A. Supra vital: living cells stained by dissociation in staining fluid.
B. Intra vital: by injection of the dye into living organism.
2. Staining by elective solubility: this is the mechanism by which stains that
are dissolved in a solvent are more soluble in the tissue component than
they are in the solvent.
E.g.: oil red
Lipid Staining: Oil Red 0 And Hematoxylin
CONTINUE….
3. Staining by chemical: chemical reaction between tissue and colorless
solution to produce a color compound.
The reaction either:
A. True dye: dye bind directly to tissue. E.g: PAS
B. Not true dye: dye react with tissue
constituents and form a colored product. E.g:
perl’s Prussian blue

Pearls Prussian Blue- Hemosiderosis Liver - Iron stain

 CONTINUE….
4. Metallic impregnation: metallic compound (ammonical silver)
reduced to metallic state producing black deposit on tissue.
Substances that reduce ammonical silver to metallic silver directly
called argentaffin. E.g: melanin
While substances those need mordant called argyrophil. E.g:
reticulin fiber
 silver technique for melanin

Reticulin fiber stained by silver technique

 CONTINUE….

5. Staining with dyes: the largest method of staining technique,

according to the nature dyes divided into:
A.Natural dyes: produced from natural source. E.g: hematoxylin
B.Synthetic dyes: a group of organic chemical compounds.
E,g: eosin
 CONTINUE….
 Any true dye contain two groups:
A. Chromogen group: group responsible for coloration by absorption of visible
light.
B. Auxochromic group: an ionizing group convert chromogen into true dye which
increase intensity of color.
 The autochrome may be basic:
A. Basic dye: hematoxylin
B. Acidic dye: Eosin
C. Natural dye: Romanowsky dye
 METACHROMAISA
 Dyes combine with tissue and give colored compound different from the original
color of dye and color of tissue.
 Dye is known as metachromatic dye, e.g: toluidine and methylene blue.
 Methylene blue and toluidine blue are absorbed by a variety of basophilic
substrate in the tissue. Chromatin stains orthochromatically blue, but cartilage
matrix, mast cell granules metachromatically reddish purple.
 FLUORESCENT STAINING
 They have ability of converting ultraviolet light into visible light
when combined with tissue.
 We use fluorescent microscope
 E.g: fluorochrome dye and acridine orange
 PROGRESSIVE AND REGRESSIVE STAINING

 Progressive staining: this technique is one in which the different

elements in the tissue are colored in sequence at the correct time.
 E.g: mayer’s hematoxylin
 Regressive staining: this technique is one in which the tissue is first
overstained and then stained or differentiated by removing excess
stain from unwanted parts of the tissue.
 E.g: harris hematoxylin
 DIFFERENTIATION
 De-staining or differentiation: is the removal of washing out of the excess
stain until the color is retained only in the tissue components to be
studies.
 Differentiation is used only with regressive stain and can be done by :
 Washing in simple solutions include: water, alcohols
 Acid solutions e.g: picric acid
 Oxidizing agents
DIRECT AND INDIRECT STAINING
 The stains work with out adding mordant is called direct staining
E.g: methylene blue
 While the stains that need mordant is called indirect staining
E.g: hematoxylin
THANKS ANY
QUESTION ?