Development, validation and application of an enzyme immunoassay (EIA) of atriopeptin

A rapid, convenient, and sensitive enzyme immunoassay (EIA) for atriopeptin (AP) has been developed. The tracer-ligand for the assay is the 24-amino acid peptide, AP24, which has been covalently coupled to the tetrameric form of acetylcholinesterase (AChE) (EC 3.1.1.7). Tracer, unknown, and primary antibody are incubated in a 96-well microtiter plate precoated with secondary antibody. After washing, a colorimetric reaction is used to measure acetylcholinesterase activity. A direct linear correlation was obtained when comparing the conventional radioimmunoassay and the EIA by using the same primary antibody to assay: plasma samples (rat or human), HPLC column fractions, or atrial extracts. Besides being technically much less demanding and not requiring the use of the radioisotopes, the EIA is more sensitive than the radioimmunoassay and thereby lends itself to a "flash" same-day assay of samples.