Basic Immunocytochemistry for Light Microscopy

Immunocytochemistry may be defined as the identification of a cell- or tissue-bound antigen in situ , by means of a specific antibody-antigen reaction, tagged microscopically by a visible label. Successful immunocytochemistry therefore requires (1) preservation of the antigen in a form that is recognizable by the antibody, (2) a suitable antibody, and (3) an appropriate label. The basic technique was first described by Coons and colleagues (1 –3 ), who employed antibody directly labeled with a fluorescent tag to identify antigen in tissue sections. Since that time, the technique has been refined and expanded enormously. Some significant developments include the use of horseradish peroxidase (4 ) and alkaline phosphatase (5 ) as label molecules; the development of many, increasingly sensitive, multilayer detection methods; and exploitation of the strong binding between avidin and biotin in detection techniques (6 ,7 ).