Technical and Interpretative Problems with Immunofluorescence

Abstract

The publication of a paper entitled “Immunological properties of an antibody containing a fluorescent group” inaugurated use of the now rapidly expanding technique of immunofluorescence [1]. This was the first real approach to the use of a chemical attached to antibody as a tracer enabling the identification and localization of antigen [6]. Following this work, [12] localized adrenocorticotrophin in normal tissue, thus demonstrating the remarkable degree of specificity of the reagents used in the immunofluorescent technique. This set the scene for what is now a very widespread, complex subject, the localization of tissue antigens by a variety of sensitive antisera. One of the major obstacles to the widespread acceptance of this technique was the difficulty of obtaining suitable microscopes, since the main way of identifying the antibody/antigen complex linked to a fluorochrome was to excite the fluoro-chrome with UV or blue light [6]. The first commercial fluorescent microscopes were developed in 1955 and this problem has now been largely overcome, some form of immunofluorescent technique being applied in almost every laboratory throughout the world [6]. Its application to a variety of topics, including diagnostic bacteriology and the study of autoimmune disease, has established this method as one of the most important and sensitive available in biology and medicine.