The potential importance of sperm cryopreservation for aquaculture and conservation management seems still undervalued, probably because the available protocols often lead to reduced fertilization success. We experimentally compared the effectiveness of two different freezing extenders for cryopreservation of brown trout (Salmo trutta) semen, controlling for possible male and female effects. The methanol-glucose based extender that we tested was significantly more effective than a common dimethyl-sulfoxide based extender (a commercial cryopreservation kit). We then studied the effectiveness of the methanol-glucose based extender at different sperm-egg ratios and found no significant differences in fertilization ability of fresh and cryopreserved milt at a sperm-egg ratio of at least 110,000:1. We conclude that brown trout sperm cryopreserved with this extender can be used even at low sperm-egg ratios without significant effects on fertilization rates.