Helicobacter pylori infection is a crucial factor in the pathogenesis of several digestive disorders, including peptic ulcers, chronic gastritis, and gastric cancer. Moreover H. pylori induces disease-specific protein expression in gastric epithelial cells. The aim of the present study was to characterize proteins differentially expressed in H. pylori-infected gastric epithelial AGS cells. An in vitro model was established using a multiplicity of infection of 100 and evaluating the effectiveness of H. pylori infection by functional analyses. Changes in protein patterns were identified using a proteomic approach consisting of two-dimensional fluorescence difference gel electrophoresis and mass spectrometry. The expression of many proteins was found to be altered, and 28 of these were identified and classified as protein synthesis-and folding-related proteins, cytoskeleton proteins, metabolic enzymes, transcription-and translation-related proteins, angiogenesis/ metastasis-related proteins, cell communication/signal transduction-related proteins, or others (oxygen-regulated protein and oncoprotein). The expression profiles of eight of these proteins, laminin ␥-1 chain precursor, valosin-containing protein, heat shock 70-kDa protein, mitochondrial matrix protein P1, FK506-binding protein 4, Tcomplex protein 1, enolase ␣, and 14-3-3 ␤ were further examined in cancerous and paired surrounding normal tissues by immunoblot assay and immunohistochemical staining to identify molecular targets that may be involved in the pathogenesis of H. pylori-induced gastric diseases. On the basis of our results, valosin-containing protein, mitochondrial matrix protein P1, T-complex protein 1, enolase ␣, and 14-3-3