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Provided your protein is >90 pure as per 15% SDS-PAGE analysis, you should try seeding. On 28-Apr-2017 12:43 PM, "李霞" <[log in to unmask]> wrote: Dear all: My protein is a demethylase,has failed to obtain protein crystals,then adopt the method of in situ enzyme and get protein crystals,but the crystal is still very very small,how to optimize can get diffraction crystal,please? Best X. L