铝吸附甲型肝炎灭活疫苗抗原解离方法研究

A method for antigen dissociation of hepatitis A virus inactivated vaccine containing aluminium adjuvant

目的建立铝吸附甲型肝炎（甲肝）灭活疫苗中甲肝病毒（HAV）抗原的解离方法。方法选择20％二乙醇胺1．25mL、10％TritonX．1000．2mL和PBS8．55mL混合液作为解离液A；pH7．2的3％（w／v）柠檬酸缓冲液作为解离液B；20％（w／v）柠檬酸三钠为解离液C。分别检测3种解离液在不同的解离条件下对铝佐剂甲型肝炎灭活疫苗的解离情况，用ELISA法检测解离后的HAV抗原含量，计算抗原回收率，确定最佳解离液及解离条件，并验证其可重复性。结果3种解离液HAV抗原解离试验显示，解离液C在室温（20~28℃）条件下解离1h，HAV抗原的回收率最高，可达95％及以上，重复试验的变异系数均〈10％。结论20％（w／v）柠檬酸三钠可作为铝佐剂甲肝灭活疫苗的首选抗原解离剂，该解离剂配方简单，操作简便，结果重复性好，准确度高。

Objective To develop a method for antigen dissociation and content determination of hepatitis A virus （HAV） inactivated vaccine containing aluminium adjuvant. Methods Buffer A： 20% diethanolamine 1.25 mL, 10% Triton X-100 0.2 mL, and PBS 8.55 mL; buffer B： 3%（w/v） citric acid with pH=7.2; buffer C： 20%（w/v） trisodium citrate. The above 3 buffers were used to dissociate HAV inactivated vaccine containing aluminium adjuvant in various conditions. The content of antigen of those resultant samples was determined by ELISA, and then the recovery rate was calculated to determine the best buffer and conditions of dissociation. The repeatability of established method was verified. Results Dissociation test showed that when HAy inactivated vaccine containing aluminium adjuvant was treated with buffer C at 20-28 ℃ for 1 hour, the recovery rate was the highest of 95% or above. Coefficient of variation of repeated trials was less than 10%. Conclusions 20%（w/v） trisodium citrate is the best buffer for dissociating antigen from HAV inactivated vaccine containing aluminium adjuvant. The buffer is easy to prepare and operate with good reproducibility and high accuracy.