C群脑膜炎球菌多糖结合疫苗制备工艺的优化

Optimization of procedure for preparation of group C meningococcal polysaccharide-tetanus toxoid conjugate vaccine

目的优化C群脑膜炎球菌多糖-破伤风类毒素(Tetanus toxoid,TT)结合疫苗的制备工艺。方法分别采用衍化的降解多糖与TT结合、衍化TT与降解多糖结合和1-氰基-4-二甲基氨基吡啶.四氟化硼(CDAP)活化降解多糖后衍化再与TT结合的方法制备C群脑膜炎球菌多糖蛋白结合物PSAH-TT、PS-AHTT和PSCDAPAH-TT,并进行生化指标和抗原性检测,与溴化氰(CNBr)活化多糖衍化与TT的结合物PSCNBrAH-TT进行免疫原性比较,确定最适制备工艺,并制备2批结合疫苗,与市售疫苗进行免疫原性比较。结果 3种方法制备的多糖蛋白结合物生化指标存在差异,以PSCDAPAH-TT收率最高,且均保持了多糖的抗原性及免疫原性,并有免疫加强效应;PSCDAPAH-TT免疫2针后抗体GMT水平高于PSAH-TT和PS-AHTT,差异有统计学意义(P<0.05),3针后抗体GMT水平仍高于PSAH-TT和PS-AHTT,但差异无统计学意义(P>0.05),PSCDAPAH-TT免疫2针和3针后抗体GMT水平均高于PSCNBrAH-TT,且2针后差异有统计学意义(P<0.05),3针后差异也无统计学意义(P>0.05),但制备的2批PSCDAPAH-TT疫苗免疫2针和3针后的抗体GMT水平均明显高于市售疫苗,差异有统计学意义(P<0.05)。结论经CDAP活化降解多糖后衍化再与载体蛋白结合制备的疫苗具有较好的免疫原性。

Objective To optimize the procedure for preparation of group C meningococcal polysaccharide（GCMP）-tetanus toxoid（TT） conjugate vaccine.Methods GCMP-TT conjugates were prepared by binding degraded GCMP derivative to TT（PSAH-TT）,TT derivative to degraded GCMP（PS-AHTT）,and degraded GCMP after activation with 1-cyano-4（dimethylamino）-pyridinium tetrafluoroborate（CDAP） and derivation to TT（PSCDAPAH-TT）,respectively,then determined for biochemical indexes and antigenicity.The immunogenicity of prepared conjugates was compared with that of conjugate PSCNBrAH-TT prepared by binding CNBr-activated GCMP derivative to TT,based on which the preparation procedure was optimized.Two batches of conjugate vaccine were prepared by the optimized procedure,of which the immunogenicity was compared with those of commercial conjugate vaccines.Results The conjugates prepared by three methods maintained the antigenicity and immunogenicity of GCMP and showed immunopotentiation,while showed difference in biochemical indexes,of which the recovery rate of PSCDAPAH-TT was the highest.The GMT of antibody induced by 2 doses of PSCDAPAH-TT was significantly higher than those by 2 doses of PSAH-TT and PS-AHTT（P〈0.05）.However,after 3 doses were given,the GMT induced by PSCDAPAH-TT was still higher than those by PSAH-TT and PS-AHTT,which showed no significant difference（P〉0.05）.The GMTs of antibody induced by 2 doses of PSCDAPAH-TT were significantly higher,while those by 3 doses were insignificantly higher,than those by PSCNBrAH-TT（P〈0.05）.Both the GMTs of antibody induced by two batches of PSCDAPAH-TT were significantly higher than those by commercial conjugate vaccines（P〈0.05）.Conclusion The procedure for preparation of conjugate vaccine was optimized as follows： GCMP was activated with CDAP then derived and bound to carrier protein,by which the prepared conjugate showed high immunogenicity.