Introduction to Histology 1.

Fixation
- Begins immediately after death
Levels of Cellular Organization - Prevent alterations in their structure
1. Organism through decomposition
2. Organ System o (1) Avoid tissue destruction by
3. Organ digestive enzymes (autolysis) or
4. Tissue through bacterial degradation
5. Cell o (2) Terminate cell metabolism
o (3) Preserve the structure and
Histology molecular composition
o (4) Kill pathogenic
- Study of tissues, fxn and arrangement to
microorganisms (bacteria, fungi,
constitute an organ
viruses)
Tissue o (5) Hardens tissue by cross-
linking or denaturing proteins
- Group of cells specialized to carry on an - Commonly used fixative:
interrelated fxn and their associated FORMALIN/FORMALDEHYDE
extracellular matrix Electron Microscope o Glutaraldehyde
o Tissue= cells + extracellular matrix (preserve lipids) o Osmium tetroxide/tetraoxide
- Made up of interacting components: cells and
extracellular matrix
o Ex. Bone, Cartilage, Connective tisse
2. Decalcification (!!!)
Extracellular matrix - Only done in specimens such a bones
and calcifies tissue
- Composed of many kinds of: - Decalcification agent: NITRIC ACID
o Ground substance (HNO3)
o Fibers
- Provides support to the cells, transport 3. Dehydration
nutrients and eliminate wastes - Successively bathing the specimen in
mixture of ethanol and water from 70%
Note: --- Cells produce extracellular matrix
to 100% (increasing concentration of
--- Content of the extracellular matrix may affect alcohol)
the fxn of the cell - Alcohol will remove water out of tissue
- Dehydrating agent: ETHANOL

4. Clearing
Preparation of tissue for study
- Remove excess alcohol
- Thin and translucent histological sections - Makes the tissue translucent
- Clearing agent: XYLENE/TOLUENE
Histology Laboratory

- Fixation -> Dehydration -> Clearing -> Infiltration 5. Infiltration

-> Embedding - Tissue is placed in melted paraffin wax
in an oven at 52°-60°
- Evaporation of the clearing agent

6. Blocking/Embedding/Molding
- Tissue and paraffin will harden after
removal from oven
- Plastic resins
o Makes use of plastic solution
w/c hardens tissue by cross-
linking polymers
o Eliminates the need to use oven
and paraffin; little tissue
distortion
7. Trimming
Chemical Stains
- Tissues with negative charges/acids are
readily stained with basic dyes=
basophilic
- Nucleic acids=nucleus
- Tissues with positive charges/basic are
stained with acidic dyes= acidophilic
- Mitochondria, collagen, cytoplasm
10. Mounting
- Placing at sections of slide with
adhesive such as pinene or acrylic resins
- CANADA BALSAM

8. Cutting and Sectioning/Microtomy  Feulgen reaction- reveal DNA as a component of

- Trimmed into appropriate sized blocks chromatin and chromosomes
- Cutting is removal of excess paraffins  Periodic Acid Schiff- detect carbohydrates or
??? polysaccharides
- Block is then mounted in microtome  Sudan Black- lipids
and cut with a steel knife
- Sectioning is done with the aid of Frozen Sections
microtome (2-3 micrometers)
- Fixation is done by rapid freezing
o Compressed CO2
9. Staining
o Liquid nitrogen
- Since paraffin is colorless, staining is a
must - Sectioning is done through cryostat
- Before staining, the following should be - Staining: METHYLENE BLUE
done: - Method is rapid
o Removal of paraffin by xylol or - Routinely done is hospital to study specimens
during surgery
toluol
- Lipids and enzymes best preserved in this
o Rehydration of tissue by
method
descending concentration of
alcohol
o Most common stain in
histology: H&E (HEMATOXYLIN
AND EOSIN)
 Hematoxylin- basic dye;
usually stains nucleus
and RNA-containing
portion of cytoplasm
 Eosin- acidic dye-
usually cytoplasmic
components and
collagen
- Most dyes act acidic/basic compounds