Effect of papain digestion on the specificity of fluorescein-labeled immunoglobulins

Abstract

During initial studies, we found that many fluorescein isothiocyanate-labeled anti-immunoglobulin conjugates were unstable and tended to aggregate and precipitate when used for indirect immunofluorescence microscopy. In some instances, the precipitate was extensive enough to interfere with interpretation of the test results. Attempts to resolve this problem resulted in a procedure by which such conjugates were digested with papain to Fab and Fc fragments before use. Aggregation and precipitation were prevented, while desired antibody activity was retained. Digestion with papain also reduced the diffuse background fluorescence (commonly referred to as nonspecific fluorescence or staining) that is often associated with conjugates before they are sorbed with tissue powders or chromatographed to remove highly labeled immunoglobulin molecules.