Six methods for separate detection of two different antigens in the same tissue section.

Abstract

Recently, Lechago, Sun, and Weinstein described a method for simultaneous visualization of two antigens in the same tissue section by combining immunoperoxidase with immunofluorescence techniques (2). In their introduction they mention other possible methods, such as double immunofluorescence and double immunoperoxidase techniques. For demonstration of two different antigens in the same tissue section, however, not less than six methods, each with several possible modifications, are available, based on three techniques; autoradiography, immunoenzyme and immunofluorescence techniques. Apart from double immunoenzyme techniques, which distinguish between two differently colored enzymatic reaction products, double immunofluorescence techniques, which d istinguish between two differently fluorescing compounds, and the combination of immunoenzyme and immunofluorescence techniques suggested by Lechago et al. (2), three techniques have been described which involve some combination with autoradiography. Autoradiography may be combined with immunofluorescence techniques as suggested by Henderson and Smithyman ( 1). Autoradiography may be combined with immunoenzyme techniques as suggested by van Rooijen and Streefkerk (6). Finally, double radioisotope autoradiographic techniques may be applied to distinguish between 3Hand radioiodine-labeled antigens (3,4) or between i25J and ‘3’I-labeled antigens (5). None of the six or modifications thereof is without disadvantages or has a general applicability. Nevertheless depending on the problem to be solved, a suitable method or modification will be available in most cases. Since individual methods and modifications are published separately and in various journals, it seemed worthwhile to note all the principles together.