Direct measurement of α‐human atrial natriuretic polypeptide in plasma by sensitive enzyme immunoassay

α‐Human atrial natriuretic polypeptide (α‐hANP) in plasma was directly measured without extraction by sensitive sandwich enzyme immunoassay. Polystyrene balls were coated with monoclonal anti‐α‐hANP (ring) IgG, specific for the N‐terminal half of the ring structure including Met‐12 residue of α‐hANP or monoclonal anti‐α‐hANP (N‐terminus) IgG, specific for the N‐terminus of α‐hANP. Rabbit anti‐α‐hANP (C‐ter‐minus) Fab' specific for the C‐terminus of α‐hANP was conjugated to horseradish peroxidase. The polystyrene ball was incubated with α‐hANP standards or plasma and, after washing, with the conjugate, and bound peroxidase activity was assayed by fluorimetry (two‐step sandwich enzyme immunoassay). The detection limit of α‐hANP was 30–90 fg (10–30 attomo1es)/tube and 0.6–2.3 ng (0.2–0.75 pmol)/L using 0.04–0.05 ml of plasma. The lower detection limit was obtained using monoclonal anti‐α‐hANP (ring) IgG 1 . Plasma hANP levels of healthy men in a supine position after an overnight fast were 24.5 ± 13.2 (SD) ng/L and tended to decrease (15.3 ± 8.5 [SD] ng/L) after intravenous administration of furosemide and subsequent one hour walking. The present sandwich enzyme immunoassay could be modified further to improve the detection limit of plasma 01‐hANP (0.2 ng/L) or to perform a less time‐consuming one‐step sandwich enzyme immunoassay without much loss of the sensitivity.