Histochemistry is the study of chemical components in cells and tissues, utilizing specific stains to visualize biochemical substances. It has applications in understanding microbial infections, healing, and the structure of oral tissues, using various fixation and staining techniques. Recent advancements include in situ hybridization and confocal laser microscopy for more precise analyses.

2. What is
histochemistry?
 It is the study of qualitative identification &
quantitative assessment of chemical
groupings within cells and tissues.
 Histochemical techniques are based on
precise chemical rationales for their ability to
identify or stain different biochemical
substances.
 Visualizing Chemicals and Enzymes in
Tissue.

3. Based on chemical reactions between
cell components and stains.
The end products of the reaction are
permanent, colored precipitates that
can be viewed under the microscope.
There are stains specific to each
component of the cell, based on the
basic or acidic nature of the dye.

4. Applications
Origin- based on the chemical
substances
Progression
Microbial infections
Healing
Repair

5. Structure and chemical
composition of oral tissues
 Epithelium and its derivatives
 Connective tissue
 Cells and fibers
 Ground substance

6. Epithelium and its derivatives
 Oral epithelium
 Epithelial components of
tooth
 Salivary glands

7. Cells and Fibers
 Fibroblasts
 Collagen (types I & III)
 Reticular fibers
 Elastic fibers
Connective tissue

8. Ground substances
 Key role is adhesion and signaling events
 Composition-Mixture of macromolecules
 Proteoglycans
 Cell surface associated macromolecules
 Extra cellular macromolecules
 Glycoproteins
 Fibronectin- fibroblasts, smooth muscle, etc.,
 Laminin- epithelial cells, basement membrane
 Chondronectin- chondrocytes
 Osteonectin- osteoblasts

9. Histochemical techniques
 Fixation procedures
 Chemical
 Formaldehyde
 Acrolein
 Gluteraldehyde
 Rossman’s fluid- glycogen, glycoproteins
 Carnoy’s mixture- nucleic acid (Fuelgen reaction)
 Paraformaldehyde- lipids
 Non-chemical
 Freeze drying- cytochrome oxidases
 Freeze fracture- electron microscopy (3D picture
of cell membranes)

10. Histochemical study of teeth
and bone
 Simultaneous fixation and decalcification with
formaldehyde or gluteraldehyde and EDTA
(ethylenediaminetetraacetic acid) for light and
electron microscopic histochemistry.
 Recently techniques have been developed for
sectioning freeze-dried, undecalcified tissues.
(eg: for studying DC).
 Bone- deproteinization with conc. Hydrazine.

11. Specific histochemical methods
Carbohydrates- PAS
Proteoglycans- Toluidine blue, Azure A, Alcian
blue.
Proteins- Dinitrofluorobenzene, Ninhydrin or
Ferric ferricyanide
Lipids- Sudan dyes (Sudan black- phospholipids)
Enzymes (For eg-Phosphatases- Gomori
method)

12. Immunohistochemistry
 Immunohistochemistry takes advantage of
antigen-antibody affinity through its ability to
identify and localize proteins of interest via
detection with labelled conjugates (fluorescent
dyes).

14. Enzyme Histochemistry
 The techniques of enzyme histochemistry, which relate
structure and function, can be used to locate many enzymes,
including acid phosphatase, dehydrogenases, and
peroxidases.
 Enzymes indicates what is happening in the tissue

15. Histochemistry of Oral hard
tissues
Carbohydrates(PAS)
 Proteins (dinitrofluorobenzene, ninhydrin or
ferric ferricyanide)
Lipids(Sudan dyes)

16. Enzyme histochemistry
 Alkaline phosphatase- absorption
 Adenosine triphosphatase- pre absorptive
ameloblasts
 Acid phosphatase- resorption
 Esterase- matrix of bone and dentin
 Aminopeptidase- human osteoclasts
 Cytochrome oxidase- oxidation (osteoblasts,
osteoclasts)
 Succinate dehydrogenase- osteoclasts

17. Histochemistry of oral soft tissues
 Polysaccharides
 Proteins
 Lipids
 Mucins(PAS for neutral mucins & alcian blue
for acid mucins)

18. Enzyme histochemistry
 Alkaline phosphatase- endothelium,
basement membranes
 Acid phosphatase- degree of keratinization
 Esterase- Gingiva(keratinized areas),
salivary gland ducts, serous demilunes in
sublingual gland, taste buds, mast cells.
 Aminopeptidase
 Succinate dehydrogenase
and glucose-6-phosphate dehydrogenase
 β-glucuronidase- basal layer
 Cytochrome oxidase- crevicular epithelium
basal layer, ducts
of SG

19. Recent techniques
 In situ hybridization (identification
of gene or gene products)
 Laser spectroscopy (inorganic
components of calcified tissue)
 Confocal Laser scanning
microscope (3D picture)
 Radioautographic techniques
(explains uptake of chemicals)