Like gastric and intestinal mucins, the tight junction proteins called claudins can be used to determine the differentiation of gastric mucosa. We investigated the expression of claudins in gastric cancer and proposed a new claudin-based gastric cancer classification system. The expression of gastric (claudin-18) and intestinal (claudin-3 and claudin-4) claudins in non-neoplastic gastric mucosa (with intestinal metatplasia [IM], 78 cases; without IM, 88 cases) and 94 gastric cancers was analyzed immunohistochemically, as was the expression of gastric (MUC5A and MUC6) and intestinal (CD10 and MUC2) mucins. Heterogeneous expression of claudin-3, claudin-4 and claudin-18 was detected in advanced gastric cancer; however, there was no significant association between the claudins and the clinicopathological parameters. These gastric cancer tissues were also subclassified into claudin-based phenotypes: gastric claudin (G-CLDN), 28 cases (30%); intestinal claudin (I-CLDN), 41 cases (44%); and unclassified claudin (U-CLDN), 25 cases (26%). Interestingly, the U-CLDN gastric cancers had worse malignancy grades, not only in size and invasiveness but also in potential metastatic ability and patient outcome. Although the mucin-based gastric cancer classification was also assessed, no significant correlation was found between mucin production and clinicopathological parameters. These observations suggest that loss of claudin expression may enhance the grade of malignancy of gastric cancer in vivo. Classification of gastric cancers using gastric and intestinal claudins is a good biomarker for assessing the risk of poor prognosis.