Recombinant Anti-CD28 x Anti-CD3 Bispecific Antibody (Fab-VHH-Fc) requires expression of three polypeptide chains, encoding two heavy chains (Fc-containing chains) and one light chain, respectively. The Fab of anti-CD28 antibody variable domain is fused to the Fc-knob domain with a hinge region and the same as the VHH of anti-CD3 to the Fc-hole domain, or vice versa. The Fab-Fc and VHH-Fc fragments can be paired via the interaction of the Fc regions. This BsAb can improve T-cell activation. It is designed for the research of Tumors therapy.