ABSTRACT
A. H. Coons et al. introduced the use of fluorochrome-conjugated antibodies, and in 1942 introduced the fluorescein isocyanate-labeled antibodies. These labeled antibodies were used to stain tissues of mice, heavily infected by pneumococci. There are two ways of making the target antigen "visible" by use of fluorochrome-labeled antibodies: the direct and the indirect method. Both methods rely on a primary antibody. The primary antibody may be polyclonal or monoclonal, depending on the wanted specificity. The advantage of the direct method is greater simplicity in the staining procedure. Savings in time relative to the indirect method can outweigh the greater initial work if there are many samples to stain. The advantage of the indirect method is that the secondary antibody is commercially available with a wide range of conjugated molecules: fluorochromes, enzymes, metals, and radioisotopes. In well-prepared specimens using both the direct and the indirect method, the background is homogenous and no defined structures besides antibody stained bacteria are visible.
