In multiple clinical conditions including trauma and hemorrhage, reperfusion magnifies ischemic tissue damage. Ischemia induces expression of multiple neoantigens including lipid alterations which are recognized by the serum protein, β2-glycoprotein I (β2-GPI)2. During reperfusion, binding of β2-GPI by naturally occurring antibodies (Ab) results in an excessive inflammatory response which may lead to death. As β2-GPI is critical for intestinal ischemia/reperfusion (IR)-induced tissue damage and Toll-like receptor 2 (TLR2) is one of the proposed receptors for β2-GPI, we hypothesized that IR-induced intestinal damage and inflammation requires TLR2. Using TLR2−/− mice, we demonstrate that TLR2 is required for IR-induced mucosal damage, as well as complement activation and pro-inflammatory cytokine production. In response to IR, TLR2−/− mice have increased serum β2-GPI compared to wildtype mice but β2-GPI is not deposited on ischemic intestinal tissue. In addition, TLR2−/− mice also did not express other novel antigens suggesting a sequential response. Unlike other TLRs, TLR2−/− mice lacked the appropriate Ab repertoire to induce intestinal IR tissue damage or inflammation. Together, these data suggest that in addition to the inflammatory response, IR-induced injury requires TLR2 for naturally occurring Ab production.