摘要
目的针对传统半数细胞培养感染剂量(CCID_(50))法变异度高、操作繁琐的局限性,开发一种基于泊松分布原理的Ⅱ型单纯疱疹病毒(herpes simplex virus 2,HSV2)溶瘤病毒(oncolytic viruses,OVs)感染滴度精准定量方法,以提高质量控制的准确性与稳定性。方法通过优化Vero细胞培养条件(血清浓度和病毒感染后培养时间)建立稳定检测体系,分别采用传统CCID_(50)法和新型泊松分布法(基于高密度96孔微孔板,利用阴性孔数据进行最大似然估计)同步测定HSV2滴度。结果传统CCID_(50)法测得病毒滴度均值为1.75×10^(6)CCID_(50)/mL,相对标准偏差(relative standard deviation,RSD)为36.62%;而泊松分布法测得病毒滴度均值为7.94×10^(5)ifu/mL,变异度显著降低(RSD为12.59%)。结论泊松分布法通过充分整合微孔板数据以减少随机误差,显著提升了病毒滴度测定的精确性和操作效率,为OVs质量评估提供了标准化、可靠性高的解决方案,具有广阔的临床应用前景。
Objective To develop an accurate quantitative method of herpes simplex virus 2(HSV2)oncolytic viruses(OVs)viral titer based on Poisson distribution principle,aiming at the limitation of high variability and complicated operation of traditional half cell culture infectious dose method(CCID_(50)),so as to improve the accuracy and stability of quality control.Methods A robust detection system was established by optimizing Vero cell culture conditions(serum concentration and incubation time after virus infection).HSV2 titer was determined simultaneously by traditional CCID_(50)method and new Poisson distribution method(maximum likelihood estimation using negative well data based on high-density 96-well microplate).Results The mean virus titer measured by CCID_(50)method was 1.75×10^(6)CCID_(50)/mL,with relative standard deviation(RSD)of 36.62%,while that measured by Poisson distribution method was 7.94×10^(5)ifu/mL,with significantly reduced variability(RSD of 12.59%).Conclusion Poisson distribution method significantly improves the accuracy and efficiency of virus titer determination by fully integrating microplate data to reduce random error,and provides a standardized and reliable solution for OVs quality evaluation,which has broad clinical application prospects.
作者
王光裕
于雷
裴德宁
史新昌
魏长龙
周勇
梁成罡
WANG Guangyu;YU Lei;PEI Dening;SHI Xinchang;WEI Changlong;ZHOU Yong;LIANG Chenggang(National Key Laboratory of Drug Regulatory Science,National Institutes for Food and Drug Control,Beijing 100050,China)
出处
《中国生物制品学杂志》
2026年第3期326-330,共5页
Chinese Journal of Biologicals
基金
中国医学科学院中央级公益性科研院所基本科研业务费专项资金(2023-PT350-01)
药品监管科学全国重点实验室课题资金(2023SKLDRS0123)。
