Monitoring ovarian function by a solid-phase chemiluminescence immunoassay

Abstract

Abstract. Ovarian function in women may be monitored by a simple, solid-phase, chemiluminescence immunoassay for the measurement of oestrone-3-glucuronide in diluted urine. An IgG fraction of antiserum to oestrone-3-glucuronyl-6-bovine serum albumin is passively adsorbed to the walls of polystyrene tubes. Oestrone-3-glucuronyl-6-aminoethyl-ethyl-isoluminol is the labelled antigen. Daily samples of early morning urine are diluted in buffer (1:100; v/v), and 100 μl removed, in duplicate, for assay. After the binding reaction (1 h at 4°C), the solution is removed by aspiration. The antibody-bound fraction is washed once with buffer (400 μl). Sodium hydroxide (2n, 200 μl) is added and the mixture incubated for 60 min at 22°C. Luminescence is initiated by oxidation of the label with microperoxidase/hydrogen peroxide and the signal integrated for 10s. An evaluation of the method gave the following values: sensitivity of calibration curve 3.12 ± 0.75 pg/tube (mean ± sd). The intra-assay precision (CV%) was 9.52 (20 replicates; 38.4 ± 3.66 nmol/l) and 8.61 (15 replicates; 102.4 ± 8.82 nmol/l). The inter-assay precision (CV%) was 11.9 (mean of 4 pools−7.03, 23.16, 52.11 and 117.53 nmol/l over 2 months). The mean bias was −0.78% over the range 28 to 448 nmol/l and different aliquots (equivalent to 0.25 to 4 μl) of daily early morning urine gave results that were in good agreement. The concentration (nmol/l; mean ± sd) of oestrone-3-glucuronide in samples of early morning urine collected from healthy women during the early follicular, periovulatory and luteal phases of the menstrual cycle were 40.2 ± 9.9, 102.3 ± 39.4 and 84.3 ± 13.3 nmol/l, respectively. In addition, the results obtained from the analysis of EMU throughout 6 complete menstrual cycles were in good agreement with the values derived by radioimmunoassay (r = 0.94).