Multiple-reaction cycling: a method for enhancement of the immunochemical signal of monoclonal antibodies.

Most current studies using immunochemical and immunohistochemical procedures to detect antigen-antibody complexes employ some type of indirect method. Such procedures afford signal amplification because several marker-conjugate molecules can bind to each primary antibody molecule. We have observed that for monoclonal antibodies an even greater amplification can be afforded simply by performing two (or more) reaction cycles (i.e., primary antibody, secondary antibody-primary antibody, secondary antibody-etc). In the present report, we demonstrate the utility of this method for immunohistochemical (immunofluorescence) and immunochemical (ELISA: enzyme-linked immunosorbent assay) procedures employing well-characterized monoclonal antibodies directed against avian type IV (basement membrane) collagen.