IMMUNOCHEMICAL CHARACTERIZATION OF A POLYCLONAL HUMAN-ANTIBODY TO FACTOR-IX

Abstract

Inhibitors of clotting factors occurring in humans are often antibody molecules synthesized in response to exogeneous proteins used in replacement therapy. Extensive studies of inhibitors to factor VIII indicate such antibodies may be monoclonal or polyclonal in nature. To date, only 1 factor IX inhibitor was subjected to detailed immunochemical analysis, and it appears to be a monoclonal Ig[immunoglobulin]G4 .lambda. antibody. A 2nd inhibitor of factor IX was discovered in a patient with severe hemophilia B; it was subjected to immunochemical analysis. Studies on this 2nd inhibitor were done before and after an anamnestic response. Column chromatography, preparative zone electrophoresis and specific inhibitor neutralization assays using monospecific heterologous antisera to human Ig classes, subclasses and L chain types indicate that the antibody is of the IgG class and contains .kappa. and .lambda. L chains and probably all 4 IgG subclasses. The inhibitor appears to be polyclonal by immunochemical and structural criteria. Preparative isoelectric focusing of pre- and postanamnestic inhibitor samples indicates that recruitment of new clones of IgG antibody occurs as a result of anamnesis. An antibody initially restricted in Ig subclass probably became polyclonal following an anamnestic response.