CD133 has been widely investigated as the specific marker of CSCs of brain tumors, prostate cancer, melanoma and pancreatic cancer, but there are still some controversial results indicating that CD133 negative cells might also include CSCs [4, 5, 23, 24]. In our study, CD133 was generally expressed in the lumen of carcinoma glands, which was also reported by other previous research . However, besides luminal expression, cytoplasmic location was another kind of expression and previous research showed that the cytoplasmic expression of CD133 was related to metastasis and tumor progression, but this relationship was not observed in luminal expression . The GC patients with CD133 positive expression were related to poorly differentiation, and had significantly poor survival outcomes than those with CD133 negative expression [26–27]. In our study, although CD133 expression was related to well or moderate differentiation, CD133 expression was also associated with N2-3 stage, which was also similar to previous studies [28–29]. However, we did not found that CD133 was related to survival outcomes in IHC. Instead, we demonstrated that the patients with CD133 high expression had significantly worse prognosis through qPCR.