ENZYME IMMUNOMETRIC ASSAY OF THYROLIBERIN (TRH)

An enzyme immunometric assay of thyroliberin (TRH) using monoclonal an tibodies and a derivatization procedure is described. This assay, name d SPIE-IA, involves a four step procedure after chemical derivatizatio n of TRH and biological samples by diazotized APEA. Step 1: derivatize d TRH was immunocaptured by a monoclonal anti-TRH antibody coated on a 96-well microtiter plate. Step 2: after washing, derivatized TRH was cross-linked via its amino group to the wells using glutaraldehyde. St ep 3: washing and treatment with NaOH. Step 4: measurement of bound TR H using a monoclonal anti-TRH antibody labeled with acetylcholinestera se. The minimal detectable concentration was 0.1 pmol/ml: with a coeff icient of variation less than 10% in the 0.156-10 pmol/ml range. This assay is 26-foId more sensitive and more specific than the competitive enzyme immunoassay using the same monoclonal capture antibody, deriva tized TRH and TRH-acetylcholinesterase conjugate as tracer. Good corre lation was observed between SPIE-IA and a sensitive competitive enzyme immunoassay using polyclonal antibodies.