Porous carbon spheres (PCS) with different specific surface areas and porous structures were prepared by templated self-assembly method. The as-prepared PCS had uniform radically distributed channels, single cavity and dispersed dendritic pore structures respectively. The whole blood clotting time tests showed the good hemostatic properties of PCS, even better than Celox. It's probably the specific surface area made the difference over the hemostatic performance of PCS. The possible reason is that the higher specific surface area can increase the contact area between PCS and blood, promote blood coagulation rapidly. And the oxygen polar functional groups on the surface of PCS can rapidly stimulate platelets and red blood cells when in contact with blood. Both of the hemolysis rate and cell viability experiments proved that PCS had good biocompatibility. The hemostatic effect of PCS in vivo were preliminary validated on the mouse tail-cutting model and the rabbit liver injury model, and all PCS could efficiently control the caudal and hepatic hemorrhage. Especially, PCSF127 with a pore size of 3 nm and specific surface area of 335 m2 g−1 showed the best hemostatic efficiency, and it was expected to be an ideal candidate for rapid hemostasis in the battlefield or pre-hospital. Porous carbon spheres with different pore structures and specific surface areas were prepared by templated self-assembly synthesis method. The negative charges carried by the porous carbon spheres activate coagulation factor FXII to trigger a coagulation cascade reaction to complete rapid hemostasis, and the hydroxyl groups on the surfaces, as oxygen polar groups, can rapidly stimulate erythrocytes and platelets when contacting with blood, thereby having a good application prospect in the fields of rapid hemostasis and wound care. Display omitted •Porous carbon spheres (PCS) were prepared with templated self-assembly.•The microstructure made the difference over the hemostatic performance of PCS.•PCS could efficiently control the caudal and hepatic hemorrhage.