Flying-fox species worldwide are under threat of extinction. Artificial insemination (AI) has the potential to play a primary role in the conservation of endangered flying-foxes, through the genetic and reproductive management of captive colonies. Semen from surviving wild populations, or from separate captive colonies, can be utilised to maintain genetic vigour, thus preventing in-breeding in potential seed populations that can then be returned to restored habitat. The development of AI technology in flying-foxes has been hampered by the atypical reproductive biology of female Megachiroptera. Pteropids have a duplex uterus, with separate cervices, and a well-defined ovarian vascular complex that provides a counter-current exchange system between the ovary and ipsilateral uterine horn. This arrangement reduces systemic circulation of steroid reproductive hormones and makes it difficult to accurately characterise the endocrinology of the oestrous cycle; it is also consistent with the apparent lack of overt behavioural oestrus in these species. Low concentrations of peripheral oestradiol also mean that vaginal cytology is not a strong correlate of reproductive status. If AI is to be utilised as a conservation strategy in flying-foxes, it is vital that an accurate method of oestrus detection or ovulation induction be established. The integrated examination of plasma hormones, behaviour and vaginal cytology, following direct hormonal stimulation of folliculogenesis in the ovaries, may improve the signal to noise ratio in this subtle physiological system. Such improved sensitivity may make it possible to develop an accurate method of oestrus detection. Combined with the continuing development of the remaining steps in AI, this will ensure the progress of establishing an AI protocol in flying-fox species. Pp. 128-135 in The Biology and Conservation of Australasian Bats, edited by Bradley Law, Peggy Eby, Daniel Lunney and Lindy Lumsden. Royal Zoological Society of NSW, Mosman, NSW, Australia. 2011.