Tissue preparation - Overview
The preparation of biological tissues for microscopic study requires a series of steps diagrammed in this image.
The first step in the preparation of a tissue or organ sample is chemical fixation. The goal of fixation is to preserve cell and tissue structure in a state that resembles that in the living state. The process of fixation prevents autolysis (self-digestion) by hydrolytic enzymes and provides rigidity by either cross-linking or denaturing protein molecules. The most commonly used fixative is formaldehyde.
The second step is embedding, which involves the infiltration of tissue spaces with a medium that will provide support during subsequent sectioning. The most commonly used embedding media are paraffin wax and epoxy plastics. The embedded tissue along with the medium is called the tissue block.
The third step is sectioning, which produces tissue slices sufficiently thin to allow transmission of the illumination source through the section. For light microscopy (LM), the illumination is white light; for electron microscopy (EM), the illumination is a beam of electrons. Tissue is sectioned on a microtome and sections are collected either on glass slides (LM) or mesh grids (EM). Paraffin sections for LM range from 5 to 20 microns; epoxy sections for EM range from 30 to 60 nanometers.
The fourth step is staining, which provides contrast and differentiation of the tissue structures.