ESTABLISHMENT OF AN EFFICIENT ENZYME-LINKED-IMMUNOSORBENT-ASSAY FOR THE DETERMINATION OF HUMAN CHOLINE-ACETYLTRANSFERASE

A specific and sensitive two-side enzyme-linked immunosorbent assay (s andwich-ELISA) was established for the reliable quantification of huma n brain and placental choline acetyltransferase (ChAT). In contrast to the radiometric assay developed by Fonnum, which is widely used for t he measurement of enzyme activity, the sandwich-ELISA particularly rec ognized inactivated forms of the antigen. In the assay, affinity-purif ied polyclonal synthetic peptide antibodies adsorbed to the polystyren e surface of the microtiter plate were employed as capture reagent. Ba sed on standard peroxidase protocols, immobilized ChAT was detected us ing monoclonal antibodies raised against human placental ChAT. By use of this ELISA, ChAT was determined at various purification stages of t he enzyme, in body fluids, during recovery experiments and in sera of patients with severe brain damage.