Use of blocking ELISA additivity test and antibody-antibody competition technique for analysis of recognition ability of monoclonal anti-gonadotropin releasing hormone antibodies.

ABSTRACT

Monoclonal anti-GnRH antibodies reacting to the heptapeptide 4-10 (H-Ser-Tyr-Gly-Leu-Arg-Pro-Gly-OH) were isolated by affinity chromatography using Sepharose 4B-heptapeptide (4-10) column. The ELISA additivity test and antibody-antibody competition techniques were used to study whether the affinity purified MoAb (A-MoAb) fraction recognize the sequence or the conformation of the native hormone. All four A-MoAbs, P862, P778, P764 and P813, were able to recognize the common epitope and did not allow to bind the conventional anti-GnRH antibodies (CoAbs) indicating that the CoAbs were conformation specific. Similarly in antibody-antibody competition technique, all A-MoAbs were able to compete with CoAbs, indicating that MoAbs were generated against the conformation of GnRH involving the entire molecule.