The immunological response in the regional lymph nodes (LN) of gastric cancer patients who had metastasis was studied by investigating both the degree of metastasis and the maturation status of dendritic cells (DCs). A total of 732 LNs was obtained from 29 gastric cancer patients, including 25 patients whose LNs were pathological metastasis-negative and 4 patients with metastasis-positive LNs. Micrometastasis (MM) in the LNs was analyzed by immunohistochemical staining of cytokeratin (CK) (IHC-MM) and by amplifying CEA and CK19 mRNA by quantitative real-time RT-PCR (PCR-MM). Distribution and density of mature DCs were evaluated in 119 LNs from pathological metastasis-positive cases by examining CD83 expression immunohistochemically. Then, following the examination of PCR-MM, immunological responses were analyzed by amplifying CD83, CD86, CD1a and IFNγ mRNA by quantitative real-time RT-PCR in 613 LNs from 25 pathological metastasis-negative cases. Among 119 LNs from 4 patients with LN metastasis, 20 LNs had histological metastasis and 6 histological metastasis-negative LNs had IHC-MM. The distribution and density of mature DCs were identical in the LNs regardless of metastasis status. Among 613 LNs from 25 patients without histological LN metastasis, 15 LNs (2.45%) from 6 cases (24%) were PCR-MM positive. The expression levels of CD83 and CD86 mRNA were significantly higher in the 15 PCR-MM-positive LNs than in the 598 PCR-MM-negative LNs. The expression levels of CD83 and CD86 mRNA in 136 PCR-MM-negative LNs from 6 PCR-MM-positive cases were also higher than those in 462 LNs from 19 PCR-MM-negative cases. In gastric cancer patients, activation of the immune response by maturation of DCs is wide-spread in regional lymph nodes at a stage before metastasis is detected histologically, including detection by IHC-MM.