Quantitation of Group-specificaAntigen in Hepatitis B Vaccines by Anti-HBs/aMonoclonal Antibody

Balb/c mice were immunized with aluminium hydroxide [alum, Al(OH)3]-adjuvanted hepatitis B (HB) vaccines of subtypesadr,ayworadw. Spleen cells from the immune animals were fused with SP2/O cells. Eight hybridoma clones producing antibodies specific or HB surface antigen (HBsAg) were selected. Monoclonal antibodies (mAbs) of four clones were specific for group-specific antigen/a, and the other of four clones were specific for subtype antigen/d,y,r, orw. The anti-HBs/amAbs were classified into three non-competitive groups. Quantitation of group-specific determinantaof HBsAg (HBsAg/a) was performed by sandwich enzyme-linked immunosorbent assay (ELISA), in which a solid phase of anti-HBs guinea-pig polyclonal antibodies (pAb), the HBsAg for testing, anti-HBs/amouse mAb and horseradish peroxidase (HRP)-conjugated anti-mouse IgG were used. The unadsorbed HBsAg was used to establish the standard curve HBsAg/a. The lower detection limits were 0·5 to 1 ng/ml of HBsAg. Methods of solubilization of alum were investigated to quantify HBsAg/ain adsorbed HB vaccines. The recovery rate was more than 60% if vaccines were prediluted. The recovery of HBsAg/ain HB vaccines produced by the same manufacturer showed the similar recovery rate, and the contents of HBsAg/ain adsorbed HB vaccines could be estimated by the recovery rate determined for adsorbed HB vaccines.