Using a split-ejaculate technique, two experiments were conducted to determine the influence of sperm exposure to seminal plasma on freeze-thaw injury. In Experiment I, 14 ejaculates from 10 Holstein bulls were held at 32°C for either 0, 20, 40, 60, 120 or 240 min followed by dilution in egg yolk-citrate. All treatments were maintained at 32°C for 240 min post-collection, at which time semen was cooled to 5°C, glycerolated, and then frozen in .5-ml French Straws using N₂ vapor. Experiment II, using 18 ejaculates from 10 bulls, was conducted identically to Experiment I, except semen was cooled to 5°C immediately after each dilution. Semen was thawed at 5°C and incubated at 37°C. Direct counts of intact acrosomes and estimates of percent motility were recorded at 0, 2, 4, 8 and 10 hrs of incubation. In Experiment I, there was a highly significant interaction (P < .01) between holding time x ejaculates to seminal plasma with regard to acrosomal retention. Optimum exposure time ranged from 20 to 120 min and 240-min exposure was deleterious for all ejaculates (P < .05). Variation in motility was not significant among treatments. In Experiment II, holding time x ejaculates interaction was again the most significant factor (P < .01). However, 20-min exposure to seminal plasma resulted in optimum acrosomal retention post-thaw for 15 ejaculates, while 40-min exposure was optimum for 2 ejaculates and 1 ejaculate did not respond favorably to any exposure time. While degree of response to seminal plasma exposure time varied among ejaculates, 20-min exposure was not deleterious to any ejaculate. Post-thaw motility was significantly (P < .01) reduced by 240-min exposure to seminal plasma.