A prospective measurement of follicle stimulating hormone (FSH) and oestradiol between cycle days 2 and 5 was conducted to investigate the intra- and inter-cycle variability in a healthy population of women with regular menstrual intervals. Daily serum samples were obtained from 44 women for a total of 66 cycles on cycle days 2, 3, 4 and 5. FSH concentrations were consistent on all cycle days measured. Oestradiol concentrations on cycle day 2 were not different from cycle day 3, but concentrations on cycle day 4 and cycle day 5 were statistically different from both cycle day 2 and cycle day 3 by analysis of variance (P < 0.05). Evaluation of functional ovarian reserve by cycle day 3 FSH measurement has become the standard in most assisted reproductive technology programmes. The recent change in FSH standardization coupled with the inflexibility of cycle day 3 testing has led to a re-evaluation of testing protocols. Cycle day 3 appears to have emerged as a dictum because most ovulation induction protocols are initiated on cycle day 3, 4 or 5. Flexibility of sampling day can be introduced as suggested by these results. The additional information ascertained from oestradiol testing as applied to evaluation of ovarian reserve warrants further investigation.