ISCOMS重组乙型肝炎疫苗的研究

目的　提高CHO细胞表达的重组乙型肝炎病毒表面抗原 (HBsAg)的免疫原性。方法　采用透析法制备HBsAg免疫刺激复合物 (immunostimulatingcomplexes简称ISCOMS)。用放免法 (RIA)检测免疫小鼠的抗体滴度 ;标记胸腺嘧啶掺入法 (3H TdR)检测小鼠脾淋巴细胞转化率 ;CTLL 细胞株 XTT法检测IL 2活性。结果　经电镜观察ISCOMS颗粒呈蜂窝状 ,直径为 30～ 4 0nm左右。SDS PAGE分析 ,制备的ISCOMS颗粒HBsAg蛋白均由P2 3、GP2 7和GP30组成。用Bradfords法测定ISCOMS蛋白回收率为 4 1 5 0 %。免疫NIH小鼠结果显示ISCOMS重组乙肝疫苗和铝佐剂疫苗的ED50 分别为 10 3 2 2 5 5和 2 5 80 6 4 ;ISCOMS组的相对效力是铝佐剂组的 4倍。刺激指数 (SI)分别为 6 78± 2 32和 3 12± 0 5 4。IL 2水平分别是 10 2IU ml± 2 12IU ml和 4 95IU ml± 0 86IU ml。结论　ISCOMS重组乙型肝炎疫苗稳定性好 ,免疫原性强 ,抗体产生早 ,滴度高 ,并且能激起细胞免疫 ,有可能发展成为新一代疫苗。

Preparation of ISCOMS as A Recombinant Hepatitis B Vaccine

Objective To improve the immunogenicity of recombinant HBsAg expressed in CHO cells.Methods Prepare the immunostimulating complexes (ISCOMS) of HBsAg by dialysis and immunize mice with the complexes.Determine the antibody titer, spleen lymphoyte transformation rate and IL-2 activity of the immunized mice by RIA,~3H-TdR incorporation and IL-2 dependent CTLL-2 cell line/XTT method respectively.Results Under electron microscope, ISCOMS particles, each at a diameter of 30-40 nm,showed cellular structures. SDS-PAGE proved that the HBsAg protein of ISCOMS consisted of P23,GP27 and GP30. The recovery rate of ISCOMS protein determined by Bradfords method was 41.50 %.The ED_(50) induced in mice by the ISCOMS and HB vaccine with aluminium hydroxide adjuvant were 103.225 5 and 25.806 4 respectively. The relative efficiency of ISCOMS was 4 times higher than that of HB vaccine with aluminium hydroxide adjuvant. The stimulating indexes of ISCOMS and HB vaccine with aluminium hydroxide adjuvant were 6.78±2.23 and 3.12±0.54,and the IL-2 levels induced by them were (10.2±2.12) IU/ml and (4.95±0.86) IU/ml, respectively.Conclusion The ISCOMS showed good stability and immunogenicity,and may be the candidate of a novel HB vaccine.