Double Label Immunohistochemistry on Tissue Sections Using Alkaline Phosphatase and Peroxidase Conjugates

One of the most common methods in immunohistochemistry involves the use of an antibody to the antigen of interest detected indirectly with an enzymelabeled antispecies secondary antibody. The enzyme catalyzes the formation of a colored insoluble reaction product at the antigen site. It is possible, with careful choice of reagents, to label two antigens simultaneously, resulting in two different colored reaction products (1 ). Cells or tissue sections can also be double labeled with two antispecies secondary antibodies carrying different fluorochromes (see Chapter 34), or by using suitable antibodies conjugated to different sizes of colloidal gold (see Chapter 32).