Histopathology
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Histopathology, the microscopic study of diseased tissue, is an important tool of anatomic pathology since accurate diagnosis of tumors and other diseases usually requires histopathological examination of samples.
Histopathological examination of tissues starts with surgery, biopsy or autopsy. The tissues are then fixed in a fixative, a process that stabilizes the tissues to prevent decay. The most common fixative is formalin (10% formaldehyde in water). The samples are then immersed in multiple baths of progressively more concentrated ethanol, to dehydrate the tissue, followed by toluene, and finally hot liquid paraffin. During this 12 to 16 hour process, paraffin will replace the water. Soft, moist tissues are turned into a hard paraffin block, a process known as embedding. This allows the sectioning of tissues into very thin (5 - 7 micrometer) sections using a microtome. These slices, thinner than the average cell, are then layered on a glass slide for staining.
A 5 to 7 micrometer slice of most tissues is almost completely transparent with very little visible detail. To see the tissue under a microscope, the sections are stained with one or more pigments. Hematoxylin and eosin are among the most commonly used stains in histopathology. Hematoxylin colors nuclei blue, eosin colors the cytoplasm pink. Other compound used to color tissue sections include saffron, silver salts and numerous artificial dyes originally developed to stain cloth fibers. The science of tissue staining is called histochemistry.
Recently, antibodies are used to stain specific proteins: this is called immunohistochemistry. This technique has greatly increased the ability to identify categories of cells under a microscope. Other advanced techniques include in situ hybridization to identify specific DNA or RNA molecules, and confocal microscopy. Digital cameras are increasingly used to capture histopathological images.