A 'dot-immunobinding assay' on nitrocellulose membrane for the determination of the immunoglobulin class of mouse monoclonal antibodies.

A rapid and convenient procedure for the determination of the immunoglobulin class of mouse monoclonal antibodies is described. Hybridoma supernatants or other dilute solutions of monoclonal antibodies are spotted onto nitrocellulose membrane strips, and each strip is incubated with a different rabbit antiserum specific for a mouse Ig class. The strips are then incubated with protein A conjugated to peroxidase, and finally with the peroxidase substrate 4-chloro-1-naphthol. A positive reaction produces a dark blue spot. The procedure is sensitive, economical, fast, and safe. Many monoclonal antibodies can be tested at one time, and most of the reagents can be re-used. The procedure has been applied to a new series of monoclonal antibodies recognizing plasma membrane antigens of rat thymocytes.