Abstract: The medium in which prehatching Xenopus laevis embryos were cultured (hatching medium) can solubilize vitelline envelope (VE) of dejellied uterine eggs and dimethyl casein.Western blotting analyses with antibodies against hatching enzyme revealed the presence of 60 kD,and occasionally 40 kD molecules in hatching medium.Results on VE-solubilizing activity assay indicated that the fractions containing 60 kD molecules exhibited strong solubilizing activity but those containing 40 kD alone did not,although both fractions exhibited the same level of proteolytic activities.However,solubilization of VE was obtained when the 40 kD fractions were mixed with an extremely low concentration of 60 kD fractions that can not solubilize VE by itself,or were applied to the VE-that had been pretreated with a low concentration of 60 kD fractions.We proposed that recognition and/or processing of VE by CUB repeats in 60 kD molecules made VEs solubilization possible by 40 kD molecules.