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In this short communication, a novel protocol for resolubilization of TCA-precipitated plant proteins for 2-DE is described.Guanidine hydrochloride (Gdn-HC1) is used as an intermediate for protein solubilization in which proteins are reduced and alkylated with tributylphosphane (TBP) and 2-vinylpyridine (2-VP).The blocking of-SH groups at Cys residues can greatly improve the solubility of TCA-precipitated proteins and obtain more high-quality protein spots in the 2-DE gel.This protocol is compatible with silver stain and MS identification.