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The aim of this study was to test the influence of postthaw storage time on sperm motility parameters of brook trout (n = 9). Furthermore, we examined the effect of sperm-to-egg ratios of 300,000:1 and 600,000:1 on fertility of postthaw, cryopreserved, brook trout sperm. The application of a cryopreservation procedure produced very high postthaw sperm motility (56.8 ± 4.0%). The cryopreserved sperm of brook trout could be stored up to 60 minutes without loss of the percentage of sperm motility (52.0 ± 9.0%). The fertilization capacity of brook trout postthaw sperm was comparable with the fertilization rate of fresh semen at a sperm-to-egg ratio as low as 300,000:1 (42.4 ± 14.0% and 36.5 ± 11.0% for eyed and hatched stages, respectively). The possibility of postthaw semen storage for the prolonged time and the obtainment of high fertilization rate at low sperm-to-egg ratio can lead to the significant improvement of brook trout semen cryopreservation procedure. [ABSTRACT FROM AUTHOR]