Evaluation of antibody- and antigen-coated enzymeimmunoassays for measuring oestrone-3-glucuronide concentrations in urine.

A monoclonal antibody to oestrone-3-glucuronide (OG) was generated and incorporated into antigen- and antibody-coated competitive enzymeimmunoassays (EIAs) using OG-, 6-ketoestrone-6-O-carboxymethyl-oxime (OCMO) and oestrone-3-hemisuccinate (OHS) as the steroid coating antigens or 'tracers' in each format respectively. In the coated-antigen format, standard curves with the lowest mean values (pg/well) for sensitivity (1.1 +/- 0.1), mid-point (ED50; 8.2 +/- 0.7) and high-point (ED20; 31 +/- 2) were obtained using OCMO coupled to bovine serum albumin (BSA) as the coating antigen, and horseradish peroxidase (HRP) as the enzyme label coupled directly to the monoclonal antibody. Standard curves generated using enzyme-labelled OG, OCMO and OHS as 'tracers' in the antibody-coated EIA format were all similar, but had higher mean sensitivity, ED50 and ED20 values than those obtained in the optimal coated-antigen format. In both EIA formats alkaline phosphatase (AP) was found to be inferior to HRP as an enzyme label. Measurement of OG concentrations in early morning urine samples from women with natural, regular menstrual cycles, using the antigen-coated EIA, demonstrated the characteristic elevation in OG concentrations associated with the onset of the urinary LH surge. This technically straightforward and robust antigen-coated EIA may be of interest to laboratories with a requirement to measure OG concentrations in urine, or other biological samples.