Rat pituitary monolayer bioassays were used to compare gonadotrophin surge-attenuating factor (GnSAF) bioactivity in follicular fluid from 12 follicles in 10 spontaneously cycling women with that in pooled follicular fluid from women undergoing ovulation induction. Expressed as ED50s (μl follicular fluid/well producing 50% of maximal effect), GnSAF bioactivity was detectable in all spontaneous follicular fluid samples (1.4-33.3 μl/well) and in follicular fluid from women undergoing ovulation induction (6.8 μl/well). This GnSAF bioactivity was unaffected by pre-incubation with an inhibin antibody. When the data were grouped according to whether the recovered oocytes fertilized in vitro or not, the fertilized group contained significantly greater GnSAF bioactivity than the unfertilized group (5.3 ± 1.1 and 14.1 ± 2.6 μl/well respectively, P < 0.05). While both inhibin bioactivity (9.7 ± 1.4 and 28.9 ± 12.1 μl/well) and immunoreactivity (36.8 ± 2.2 and 21.0 ± 3.0 and ng/ml) were also greater (P < 0.01) in the fertilized compared with the unfertilized groups respectively, there were no other significant differences between the two groups. We conclude that GnSAF is found in follicular fluid from spontaneously cycling women, supporting in-vivo evidence for the involvement of GnSAF in feedback control of the ovary-pituitary axis. © 1995 Oxford University Press.