Introduction to Histology and Staining Techniques

How many steps are involved in studying histology with a microtome?

6: fixation, dehydration, clearing, infiltration, embedding and trimming

Fixation

Preserves tissue structure by using cross-linking proteins.

Dehydration

Removes water using increasing alcohol concentrations.

Clearing

Removes alcohol with miscible organic solvents. (both alcohol and parafin are miscible)

Infiltration

Tissue saturates with melted paraffin, until complete infiltration with the substance.

Embedding

Paraffin-infiltrated tissue is molded and hardened.

Trimming

Paraffin block is cut for microtome sectioning (slicing).

What does staining do?

Enhances visibility of tissue and cell structures.

What happens withouts staninig?

Unstaid tissue and cells are difficult to examine.

What are the two types of dies?

Basic (+) and Acidic (-)

Basic Dyes (+)

Stain anionic (-) components, like nucleic acids.

Acidic Dyes (-)

Stain cationic components, like ionized proteins.

Hematoxylin is what type of dye?

Basic dye staining anionic structures.

Eosin is what type of dye?

Acidic dye staining cationic structures.

Hematoxylin stains what color?

Stains anionic tissues purple

Tissues that stain with basic dyes are said to be what?

Basophilic

What type of cell structures stain well with hematoxylin?

Those with acids:

Nucleic acids (nucleus contains DNA).

Glycosaminoglycans

Eosin stains what color?

Cationic structures, Pink

Tissues that stain with acidic dyes (eosin) are said to be what?

Acidophilic

What cell structures stain well with eosin?

Those with basic components:

Mitochondria

Secretory granules

Collagen

Hematoxylin & Eosin (H&E)

Common histology dye combination for tissue staining.

Are acidophilic tissues composed of acids?

NO

Basophilic

Structures that stain well with basic dyes.

Acidophilic

Structures that stain well with acidic dyes.

Periodic acid - Schiff (PAS) is what?

a special staining technique

What does Periodic acid - Schiff (PAS) stain?

Stains carbohydrates dark purple or magenta.

Example of a structure that stains with PAS?

Goblet cells: secretory granules rich in carbs are

PAS positive.

What is Sudan Black?

A lipid-soluble dye staining lipids black.

Adipose Cells

Cells rich in lipids, stained by Sudan black.

Light Microscope

Standard microscope with lower resolution and magnification.

Electron Microscope

High-resolution microscope, up to 400,000X magnification.

How does a light microscope compare to an electron microscope?

Electron microscopes provide more details and ultrastructure of the cells (organelles).

Ultrastructure

Detailed view of cellular organelles and structures.