Histopathology involves examination of stained sections of fixed tissues with a light microscope. Formalin-fixed samples submitted for histopathology are first trimmed into 2–3-mm-thick sections and placed in tissue cassettes. The tissues are then treated with a series of solutions containing increasing concentrations of ethanol to remove the water from the samples. A solvent (usually xylene) is used to displace the ethanol from the tissue, and the tissue is infiltrated with paraffin wax and submerged in a mold filled with hot liquid wax, which is subsequently cooled down to form a solid block containing the tissue. The mold is removed, and this block of wax and tissue is mounted to a microtome for sectioning. The microtome moves the block forward a set distance (the thickness of the desired section) and then down against a sharp blade to cut a thin (typically 3–5 µm) section of the paraffin block and embedded tissue. Usually, the block must be sectioned several times or “faced” initially to create an even, flat tissue surface. The thin ribbon of wax and tissue cut by the microtome is placed in a water bath and scooped up on a glass slide. The wax is removed with xylene, and the tissue is stained and covered with a coverslip. The routine stain used for histopathology is hematoxylin and eosin, but a whole array of other stains is available to highlight infectious organisms, extra- and intracellular accumulations, fibrosis, certain cell types (such as mast cells), and basement membranes.

20.1 Necropsy Samples for Histopathology

The basic set of necropsy samples for histopathology is outlined in Appendix 3. Figure 20.1 shows the approximate size of samples for histopathology. The size of the samples needs to be large enough that the person trimming the tissues can recognize the sample for appropriate orientation, but small enough to fix completely.