吸附无细胞百白破联合疫苗的细菌内毒素检查

Test for bacterial endotoxin in adsorbed diphtheria,tetanus and acellular pertussis combined vaccine

目的分析吸附无细胞百白破联合疫苗（adsorbed diphtheria-tetanus-acellular pertussis combined vaccine,DTa P）细菌内毒素检查方法的可行性。方法确定供试品最大有效稀释倍数（maximum valid dilution,MVD）,并将两个厂家的鲎试剂（λ=0.25 EU/ml）进行灵敏度复核试验后,按照《中国药典》三部（2010版）附录ⅫE细菌内毒素检查法要求,采用稀释法进行供试品干扰试验,确定无干扰稀释倍数;采用凝胶限量试验对13批DTa P进行细菌内毒素检测,并采用动态显色法定量检测其中6批DTa P细菌内毒素含量。结果供试品MVD为800倍;两个厂家鲎试剂灵敏度均在0.125-0.5（0.5-2λ）范围内,符合《中国药典》三部（2010版）规定的标准。将DTa P稀释400倍可排除干扰。13批DTa P凝胶限量试验结果均〈12 EU/剂,其中6批DTa P动态显色法结果均〈2.5 EU/剂,相同6批疫苗两种检测方法的结果趋势一致。结论凝胶限量试验与动态显色法均可用于DTa P细菌内毒素检查,但凝胶限量试验更经济,操作更简便,适用范围更广。

Objective To investigate the feasibility of test for bacterial endotoxin in adsorbed diphtheria,tetanus and acellular pertussis（DTa P） combined vaccine. Methods The maximum valid dilution（MVD） of test sample was determined,and checked for sensitivity by the limulus reagents manufactured by two manufacturers（λ = 0. 25 EU / ml）,then subjected to interference test by dilution method to determine the dilution factor without interference according to the requirements in Appendix Ⅻ E of Chinese Pharmacopoeia（Volume Ⅲ,2010 edition）. Thirteen batches of DTa P were tested for bacterial endotoxin by gel-clot limit test,while the bacterial endotoxin contents in six batches of DTa P were determined by kinetic chromogenic assay. Results The MVD of test sample was 800. Both the sensitivities of limulus reagents by two manufacturers were within the range of 0. 125 - 0. 5（0. 5 - 2 λ）,which met the requirements in Chinese Pharmacopoeia（Volume Ⅲ,2010 edition）. DTa P was diluted by 400 folds to overcome the interference. The bacterial endotoxin contents in thirteen batches of DTa P determined by gel-clot limit test were less than 12 EU / dose,while those in six batches by kinetic chromogenic assay were less than 2. 5 EU / dose. The tendencies of determination results of six batches of vaccine by two methods were in agreement. Conclusion Both gel-clot limit test and kinetic chromogenic assay may be used for the bacterial endotoxin test in DTa P. However,gel-clot limit test was more economic and simple,and worthy of popularization.