In order to elucidate the mechanisms of let-7a down-regulation in pathogenesy of gastric carcinoma, proteins associated with the function of let-7a were detected in high throughout screening. The cell line of SGC-7901 stablely overexpressing let-7a was successfully established by gene clone. Two-dimensional gel electrophoresis (2-DE) was used to separate the total proteins of SGC-7901/let-7a, SGC-7901/EV and SGC-7901, and PDQuest software was applied to analyze 2-DE images. Ten differential protein spots were identified by MALDI-TOF-MS, and they may be the proteins associated with let-7a function. The overexpressed proteins included antioxidant protein 2, insulin-like growth factor binding protein 2, protein disulfide isomerase A2, C-1-tetrahydrofolate synthase, cyclin-dependent kinase inhibitor1 and Rho-GTPase activating protein 4. The underexpressed proteins were consisted of S-phase kinase-associated protein 2, platelet membrane glycoprotein, fibronectin and Cks1 protein. Furthermore, the differential expression levels of the partial proteins (CDKN1, Spk2 and fibronectin) were confirmed by Western blot analysis. The 10 differentially expressed proteins could be divided into groups based on their functions: signal transduction, chaperone, transcription and translation, metabolism and cytoskeleton, which were involved in cell cycle, the transcription regulation, cell adherence，cellular metabolism and so on. The data suggest that these differential proteins may be associated with the function of let-7a in gastric carcinoma, and will be valuable for further to study the mechanisms of let-7a down-regulation in pathogenesy of gastric carcinoma.