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Development and validation of a simple, sensitive enzyme immunoassay for quantification of androstenedione in bull plasma

  • Mallick, Smrutirekha (Dairy Cattle Physiology Division, National Dairy Research Institute) ;
  • Kumar, BS Bharath (Dairy Cattle Physiology Division, National Dairy Research Institute) ;
  • Prakash, BS (Division of Animal Sciences, Indian Council of Agriculture Research) ;
  • Aggrawal, Anjali (Dairy Cattle Physiology Division, National Dairy Research Institute) ;
  • Pandita, Sujata (Dairy Cattle Physiology Division, National Dairy Research Institute)
  • Received : 2014.09.03
  • Accepted : 2014.12.11
  • Published : 2015.04.30

Abstract

As an alternative to radioimmunoassay a simple and highly sensitive enzyme immunoassay (EIA) was developed and validated for androstenedione quantification in plasma of Karan Fries bulls using second antibody coating technique. The wells of the microtitreplate were coated with affinity-purified goat immunoglobulin (antirabbit IgG) that binds the hormone specific antibody. The EIA was performed to analyze androstenedione directly in $40{\mu}l$ of bull plasma. The androstenedione standards ranged from 0.20 to 200 pg/$40{\mu}l$/well and the sensitivity of the assay was 5 pg/ml plasma. Serially diluted bull plasma containing high endogenous androstenedione showed good parallelism with bovine androstenedione standard curve. Intra- and inter-assay coefficients of variation (CV) were found to be 8 and 9%, respectively. Peripheral plasma androstenedione concentrations determined in young and adult bull samples ranged between 104-990 pg/ml and 184-2040 pg/ml, respectively.

Keywords

Androstenedione;Enzyme immunoassay;Bull;Plasma;Standardisation

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