Sepsis is regarded as a gene-related disorder. Growing evidence suggests that STAT3 is a master transcriptional factor that plays an important role in inflammation. In this study, we examined whether in vivo introduction of STAT3 decoy oligodeoxynucleotides (ODNs) can provide benefits in mice with cecal ligation and puncture (CLP)-induced sepsis to assess the potential role of STAT3 in sepsis-associated organ dysfunction. Activation of STAT3 greatly increased in each of major organs after CLP in time dependent manner. We confirmed that STAT3 decoy ODNs were effectively delivered into tissues of septic mice in vivo by preparing into a complex with atelocollagen given 1 h after CLP. When STAT3 decoy ODNs were given to septic mice, abnormal production of pro-inflammatory and chemotactic cytokines was significantly reduced, histopathologic changes in lung, liver, and kidney tissues were markedly improved, and led to a significant survival advantage in mice after CLP. The STAT3 inhibitor stattic mimicked the beneficial effects of STAT3 decoy ODN transfection in septic mice. These results suggest that STAT3 is a potential therapeutic target for sepsis-associated multiple organ injury.