摘要
目的建立一种元细胞百日咳疫苗的功能性抗体检测方法，为无细胞百日咳疫苗原液及成品的生产一致性评价和效力评价提供一种便捷有效的解决方案。方法优化并使用CHO细胞簇集试验方法检测鼠免疫血清的百日咳毒素中和抗体滴度。结果确定疫苗样品以1／5人用剂量腹腔免疫20～24g、5周龄的雌性NIH小鼠10只，4周后采血分离血清，倍比稀释血清用0．1IU／ml的百日咳毒素国家参考品中和2h，加入到预先培养的CHO细胞孔中孵育48h后判定簇集结果，最终不簇集孔的血清稀释倍数的几何均值即为样品中和抗体滴度。不同工艺无细胞百日咳疫苗的中和抗体滴度有显著性差异；不同工艺无细胞百日咳疫苗改良脑腔攻击试验结果通过与不通过的产品中和抗体有明显区分度。结论百日咳毒素中和抗体法大幅减少动物使用量，CHO细胞簇集试验具备一定的重复性和精密性，可以满足不同工艺无细胞百日咳疫苗成品及原液的生产一致性和效力的监测和初步评价。
Objective To establish a functional antibody detection method for acellular pertussis vaccines in order to conveniently and effectively evaluate the production consistency and potency of acellular pertussis vaccine bulks and final products. Methods Chinese hamster ovary （ CHO ） cell clustering assay was optimized and used to measure titers of neutralizing antibodies against pertussis toxin in mouse immune serum samples. Results Vaccine samples were determined to be immunized intraperitoneally with 1/5 the human dose to ten female NIH mice （20-24 g, 5-week-old）. Four weeks after immunization, blood samples were collected to isolate serum. Serially diluted serum samples were used to neutralize 0.1 IU/ml of pertussis toxin national reference product for 2 hours. Results of clustering were determined after 48 hours of incubation in pre-cultured CHO cell wells. The geometric mean of the serum dilution of the final unclustered wells was the neutralizing antibody titer of vaccine sample. There were significant differences in the titers of neutralizing antibodies elicited by acellular pertussis vaccines prepared with different manufacturing processes. Vaccine samples succeed or failed the modified intracerebral challenge assay （MICA） were easily distinguishable by neutralizing antibodies. Conclusion The method of detecting neutralizing antibodies to pertussis toxin greatly reduces the amount of animals used in research. CHO cell clustering assay that has better repeatability and precision can be used for monitoring and initial evaluation of the consistency and potency of the bulks and final products of pertussis vaccines prepared with different manufacturing processes.
作者
卫辰
晁哲
吴燕
王丽婵
骆鹏
马霄
w Wei Chen;Chao Zhe;Wu Yah;Wang Lichan;Luo Peng;Ma Xiao(Key Laboratory of the Ministry of Health for Research on Quality and Standardization of Biotech Products,National Institutes for Food and Drug Control,Beijing 102629,China)
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2018年第7期535-541,共7页
Chinese Journal of Microbiology and Immunology
