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 (sandwich-ELISA) was established for the reliable quantification of human brain and placental choline acetyltransferase (ChAT). In contrast to the radiometric assay developed by Fonnum, which is widely used for the measurement of enzyme activity, the sandwich-ELISA particularly recognized inactivated forms of the antigen. In the assay, affinity-purified polyclonal synthetic peptide antibodies adsorbed to the polystyrene surface of the microtiter plate were employed as capture reagent. Based on standard peroxidase protocols, immobilized ChAT was detected using monoclonal antibodies raised against human placental ChAT. By use of this ELISA, ChAT was determined at various purification stages of the enzyme, in body fluids, during recovery experiments and in sera of patients with severe brain damage.