The use of biotinylated monoclonal antibodies and streptavidin affinity chromatography to isolate herpesvirus hydrophobic proteins or glycoproteins

Abstract

A streptavidin/biotin-based immunoaffinity system was optimized to isolate herpesvirus (human cytomegalovirus) immediate early proteins or late glycoproteins from crude infected cell lysates. Biotinylation of the primary antibody by biotin substitution of epsilon amino groups was superior to biotin substitution of sugar residues. Biotinylation of the primary antibody was superior to that of a secondary antibody. A biotin substitution of approximately 8 M biotin/M antibody allowed for maximal recovery of viral antigens. The streptavidin/biotin-based immunoaffinity system can allow for relatively pure preparations of viral antigens that may be used for functional, immunological, or structural studies.