Use of enzyme-immunoassay for oestradiol-17β and progesterone quantification in canine serum

The objective of this investigation was to develop and evaluate competitive inhibition-enzyme-immunoassays for canine serum oestradiol-17β (E2) and progesterone (P4) quantification. Sera from 56 healthy bitches at various stages of oestrus cycle and pregnancy were tested. For E2 measurement, each sample (0.4 ml) was extracted with diethyl ether and after solvent evaporation the resultant hormone was reconstituted to one-fifth of the original sample volume in aqueous buffer. Each reconstitute (30 μl) was assayed for E2 to estimate respective serum concentration. For P4, each sample (10 μl) was directly assayed without extraction. The classic cyclic hormonal pattern during the oestrus cycle of the bitch was observed. The brief, sharp dominance of E2 during the follicular phase was followed by the long-lasting dominance of P4 during the luteal phase (late oestrus, dioestrus or pregnancy). During the anoestrus phase both hormones were found at basal levels, with the exception of E2 during late anoestrus which appeared to be rising. Both assays had acceptable specificity (cross-reactions≤10%), precision (coefficient of variation (C.V.)<7%) and accuracy (E2 recovery: 97%; P4 recovery: 104.7%). The sensitivity of E2 and P4 assay was 4 pg ml−1 and 0.28 ng ml−1, respectively.