Abstract: Velvet antler water- soluble protein ( VAWP) and soybean protein isolate ( SPI) were hydrolyzed by in vitro simulated gastrointestinal digestion and Alcalase to obtain different protein hydrolysates. The hydrolysis time was optimized according to the changes in the content of α- NH2. The < 3ku fractions of velvet antler water- soluble protein simulated gastrointestinal digest ( VAWP- SGD) , velvet antler water- soluble protein Alcalase hydrolysate ( VAWP-APH) , soybean protein isolate simulated gastrointestinal digest ( SPI- SGD) and soybean protein isolate Alcalase hydrolysate ( SPI- APH) were fractionated by ultrafiltration.The effect of < 3ku fractions on the proliferation of murine splenocytes were studied.Results indicated that the hydrolysis time of in vitro simulated gastrointestinal digestion were 2h with pepsin and 5h with pancreatin. The hydrolysis time by Alcalase was 4h. VAWP- SGD and SPI- SGD significantly promoted the proliferation of resting murine splenocytes ( ≥100μg / m L) .VAWP- SGD showed the highest stimulated effect on Con A- stimulated murine splenocytes proliferation, followed by VAWP- APH. SPIAPH had no obvious effect on Con A- stimulated murine splenocytes proliferation, whereas SPI- SGD inhibited the proliferation of Con A- stimulated murine splenocytes.VAWP- SGD also had the strongest stimulated effect on LPSstimulated murine splenocytes proliferation, followed by VAWP- APH, SPI- APH, and then was SPI- SGD.The above results showed that VAWP- SGD had the strongest stimulated effect on the proliferation of murine splenocytes, which suggested that VAWP- SGD was of great significance for the development of antler products and immune active peptide.