Principles and basics of immunohistochemistry

Abstract

Immunohistochemistry is a widely used technique, which visualize and observe specific substances present in cells, tissues, organs, and individuals, in many research field applying various and specific molecular recognition mechanisms that exist in the living body, so called antibody-antigen reaction. Morphology is what observes using a microscope such as light microscope, electron microscope, fluorescence microscope, confocal laser scanning microscope, and biochemistry is what detects the presence of substances in cells or tissues using immunoprecipitation, Western blotting, etc. Immunohistochemistry is a research method combining morphology and biochemistry. A brief summary of this basic step is to react with “antibody” that specifically binds to an “antigen”, and “visualize” the site where the antigen-antibody reaction has taken place and observe it microscopically. In the immunohistological steps, there are several important points. The first of which is to detect a substance (antigen) to be searched for clearly, easily and reproducibly using a “good primary antibody”. The second is the need to immobilize or fix substances (antigens) in tissues and cells. For this, fixation is performed. Fixation may, on the one hand, lead to a change in the molecular structure of the antigen, resulting in suppression of the antigen-antibody reaction. It is therefore necessary to overcome the reciprocity problem of fixation and immunostaining. The third is “visualization”. It must not only be visible, but must be able to be specific and clear. It is very important to be able to discriminate the correct immunohistochemical reaction while being aware of these points, and not to make a judgment on detection as a result of false detection. In addition, control experiments for that purpose are always important points to be aware of.