On the basis of evidence suggesting a maternal involvement in the determination of the sex of the offspring, we took ova at the point of ovulation from crossbred heifers, fertilised them, and established the sex of the embryos. At the same time we took individual-matched samples of follicular fluid from each follicle of origin, and measured the levels of testosterone and oestradiol, blind to the sex of the embryo. We found no effect of oestradiol on sex in either primary or subordinate follicles. But bovine ova from subordinate follicles that had follicular fluid with a high concentration of testosterone (in vivo) were later more likely to be fertilised by a Y-chromosome-bearing spermatozoon (in vitro). These, along with similar results from other researchers, suggest that further study of the relationship between mammalian follicular hormones at the time of conception and subsequent sex of offspring, may help resolve some of the problems associated with theories of adaptive control of the sex ratio in mammals. J. Exp. Zool. 303A:1120–1125, 2005. © 2005 Wiley-Liss, Inc.