Abstract: To study the immune regulated function of the recombinant protein 3D of FMDV for the multipleepitope recombinant antigen of FMDV, we cloned the 3D gene of FMDV type Asia 1, and the recombinant protein 3D was successfully expressed in E. coli and purified. The immune mixture of the purified the recombinant protein 3D and the recombinant antigen was developed as a ratio of 1∶2 (w/w). An immunogen was developed by emulsification of immune mixture with an equal volume of ISA 206 containing 50 μg of 3D and 100 μg of the recombinant antigen per dose. Five health guinea pigs, weighing 250300 g, were immunized submuscularlly at the rear leg with 1 mL of the immunogen each animal. At 28 days after the first immunization, all animals were booster immunized with the same dose of immunogen. Serum samples were collected at 0, 21, 28, 42 days after immunization, which were used for the detection of neutralizing antibodies and lymphocytes proliferation assay. In addition, all animal were challenged with 103 guinea pigs infective dose (GPID50) of FMDV type Asia1 at days 14 booster immunization, and potency of immunity of the immunogen was evaluated. In addition, the recombinant antigen/ISA 206 (100 μg·mL-1), the inactivated vaccine to FMDV type Asia 1 (1 mL) and the PBS/ISA 206 (1 mL) as the controls were used to inject the guinea pigs, respectively. The results showed that the recombinant protein 3D of FMDV not only significantly improve the levels of antiFMDV neutralizing antibodies elicited by the recombinant antigen, but induce higher percentage of lymphoproliferation in guinea pigs (P<0.05, t test), which were the same as that of the inactivated whole virus vaccine (P>0.05, t test). However, PBS could not elicit any levels of neutralizing antibodies and lymphoproliferation (P>0.05, t test). It’s speculated that the protein 3D of FMDV could significantly improve the potency of the multipleepitope recombinant antigen and is a very important immune regulated protein, which will contribute to design of novel FMDV vaccines.