Immunological and physiological validation of an enzyme-linked immunosorbent assay (ELISA) for the measurement of growth hormone in goat (Capra hircus) plasma

Abstract

The current study describes the optimization and validation of an enzyme-linked immunosorbent assay (ELISA) for the quantification of growth hormone (GH) in blood plasma of goats, Capra hircus. A microtiter plate based competitive assay was developed using a capture ELISA for anti-ovine GH and biotinylated ovine GH. Ovine GH standards were prepared in charcoal–dextran stripped goat plasma. The assay was optimized in terms of sensitivity, specificity and precision. The sensitivity of the assay was 50pg GH/100μl, which corresponds to 0.5ng/ml plasma. A dose–response inhibition curve resulting from serially diluted goat plasma was parallel to the standard curve using ovine GH and thus it confirmed the similar specificity of ovine GH standard and endogenous GH in goat plasma for ovine GH antibody. The precision of the assay, i.e., the intra- and inter-assay coefficients of variations (for repeatability and reproducibility, respectively) for the pooled plasma samples containing two GH concentrations (32 and 2ng/ml) were 6.72%, 11.2%, 7.1% and 12.8%, respectively. To physiologically validate the assay, goats were treated with human growth hormone-releasing factor (hGRF) and there was an immediate rise in plasma GH levels in hGRF-treated goats. This ELISA is economic, safe, quick (requires only 48h), convenient and the first competitive enzyme immunoassay described for caprine GH.