Heterologous enzyme immunoassay for serum androstenediol

Abstract

A heterologous enzyme immunoassay for serum androstenediol (Adiol: 3β, 17β-dihydroxy-androst-5-ene) was established. The combination of anti-Adiol antiserum raised in rabbit against Adiol 7- O-(carboxymethyl)oxime (Adiol 7-CMO) conjugated bovine serum albumin (Adiol 7-CMO-BSA) and Adiol 7-iminomethylcarboxylic acid conjugated alkaline phosphatase was used for the assay. The sensitivity of the heterologous assay system was superior to that of a homologous assay system in which an antibody raised in rabbit against Adiol 7-CMO-BSA and enzyme labeled antigen, Adiol 7-CMO conjugated alkaline phosphatase, were used. The minimal amount of Adiol detected was 0.4 ng ml −1 and the measurable range was from 0.4 to 150 ng ml −1. Intra-assay coefficients of variation (C.V.) were 8.6% (1.52±0.13 ng ml −1, mean±S.D., n=10) and 6.7% (13.4±0.9 ng ml −1, n=10). Inter-assay C.V. were 12.9% (1.63±0.21 ng ml −1, n=8) and 11.5% (12.2±1.4 ng ml −1, n=8). A linear relation was observed between the serum sample dilution and the Adiol concentration. For recovery study, authentic Adiol was added to serum sample (original concentration: 1.43 ng ml −1). The calculated final Adiol concentration was 2.99 ng ml −1. The recovery was 98.6% ( n=5). The Adiol concentrations in healthy subjects measured by the proposed assay (male: 1.1±0.3 ng ml −1 (mean±S.D.), range: 0.7–1.7 ng ml −1, age: 22–50, n=10; female: 0.6±0.4 ng ml −1, range: 0.2–1.6 ng ml −1, age: 23–48, n=20) were consistent with reported values.