Aim To measure and analyze the changes of plasma soluble Sema4D levels during cardiopulmonary bypass(CPB)with ELISA, and analyze possible mechanisms. Methods Recombinant extracellular Sema4D was used as antigen, and antibodies against Sema4D were used to establish ELISA method. Samples from patients during CPB were collected at three time points: before surgery, 10 min after heparinization, and 10 min after neutralization after the surgery.Soluble Sema4D levels in the plasma were measured in these samples with ELISA. Sema4D expression on the surface of T cells and platelets were detected by flow cytometry. Results Sandwich ELISA method was established to detect soluble Sema4D levels in human plasma. Soluble Sema4D levels increased during CPB and remained at high levels until the end of the surgery(P＜0.01). Compared with before surgery, percentage of Sema4D high platelets increased 10 min after heparinization in 75% patients(P＜0.05). There is no significant change of Sema4D expression on the surface of T cells during CPB.Conclusions Platelets activation and release of Sema4D may be one source of the increases of plasma soluble Sema4D during CPB. The change of plasma soluble Sema4D levels may become a prognostic indicator of CPB.