Biodegradable microspheres made from poly-lactide-co-glycolide polymers have been considered as a new delivery system for single-dose vaccine. Purified tetanus toxoid (TT) was encapsulated in poly-lactide(PLA) and poly-lactide-co-glycolide (PLGA) microparticles using a solvent evaporation method in a multiple emulsion system (water-in oil-in water). The morphology of 77-loaded microparticles was spherical and the suface of them was smooth. The particle size was in a range of 2-10. Protein loading efficiency was 68-97.8%. PLGA (85:15) microparticle showed the highest efficiency. Protein release pattern was influenced by polymer molecular weight and composition. The release rate of PLA(Mw 100,000) microsphere was higher than any other microspheres. In consequence of the hydrolysis of PLGA(50:50) microspheres, environmental pH decreased from 7.4 to 5.0. The PLA, PLGA (75:25) and PLGA (85:15) microshperes showed no significant pH change. The antigenicity or n in microshperes was assayed by indirect sandwich ELISA using equine polyclonal tetanus antitoxin for capture antibody and human polyclonal tetanus antitoxin for primary antibody. The antigenicity of TT in PLA (Mw 100,000), PLGA(50:50, Mw 100,000) and PLGA (75:25, Mw 73,300) after 30 days incubation showed 54, 40.9 and 76.7%, respectively.