Objective To study the effect of buffer capacity in up chamber on vertical streaking(VSTK) of two-dimensional electrophoresis.Methods The same buffer volume was kept in up chamber,VSTK was compared by running two SDS gels or single one in 1× Laemmli buffer(buffer),or two ones in 1.5×,2× and 3× buffer in up chamber respectively.Results VSTK appeared severely when two gels were run in 1× buffer,while disappeared when one did.When two gels were run in 1.5×buffer,VSTK only existed in the region of high molecular weight,and protein spots in the region of medium and low molecular weight were round without streaking.When two gels were run in 2× buffer,compared with 1.5× buffer,the VSTK in the region of high molecular weight was further improved,and the round protein spots in the region of medium and low molecular weight increased.VSTK disappeared completely when two gels were run in 3×buffer.Conclusion Insufficient buffer capacity in up chamber may lead to vertical streaking.