Development, Characterization and in-vitro Evaluation of HBsAg loaded PLGA microspheres stabilized with protein stabilizers

Abstract

The objective of this study is to develop a stable single-dose vaccine based on recombinant hepatitis B surface antigen (HBsAg) in PLGA microspheres in which HBsAg is stabilized using protein stabilizers and an antacid and to demonstrate its potential as mucosal adjuvant for vaccines. Preliminary studies like Surface morphology, size distribution analysis, percentage entrapment efficiency and in-vitro analysis were performed. In the entire study tetanus toxoid (TT) was used as model antigen and HBsAg was used as candidate antigen. Stability studies were carried out for the microspheres with and without stabilizers at 4’ C and 37⁰ C with RH 605%. Percentage entrapment efficiency and EIA / Protein ratio were investigated. The size distribution analysis suggested that the particle size of individual microsphere was remained same in the range of 1-10 tim. The percentage entrapment efficiency of the model antigen loaded microspheres and the candidate antigen loaded microspheres were up to 80 on 16" day and was dropped to 65 on 19" day. The integrity of the antigen was assessed by western blot analysis and was found to be unaltered up to 16 days at 37⁰C. The in vitro immunogenicity of encapsulated HBsAg was evaluated by EIA assay and the EIA/protein ratio of both the antigen was up to 0.9ñ0.2 on 16" day and was dropped to 0.7+0.2 on 19" day. To conclude the HBsAg containing PLGA microspheres were found to be stable for 18 months, 16 days and 84 days respectively at 4⁰ C, 37’ C and 25' C.