重组腺病毒疫苗插入基因表达产物鉴别方法的建立及验证  

Development and validation of a method for identification of expressed products of inserted gene in recombinant adenovirus vaccine

作　　者：孙博[1,2] 刘晓晖 马达[4] 原秀娟 闫慧 刘新涛 张海红 谷铁军[1,2] 吴婷婷 SUN Bo;LIU Xiao-hui;MA Da;YUAN Xiu-juan;YAN Hui;LIU Xin-ao;ZHANG Hai-hong;GU Tie-jun;WU Ting-ting(School of Lijfe Sciences,Jilin University,Changchun 130012,Jilin Province,China;不详)

机构地区：[1]吉林大学生命科学学院,吉林长春130012 [2]吉林大学艾滋病疫苗国家工程实验室,吉林长春130012 [3]吉林省产品质量监督检验院,吉林长春130022 [4]长春百克生物科技股份公司,吉林长春130103 [5]长春安沃高新生物制药有限公司,吉林长春130012

出　　处：《中国生物制品学杂志》2022年第9期1102-1106,1117,共6页Chinese Journal of Biologicals

基　　金：吉林省科技发展计划-双靶点新型肿瘤治疗性疫苗临床前和临床研究(20180201001YY)。

摘　　要：目的建立重组腺病毒疫苗插入基因表达产物的鉴别方法,并进行验证。方法采用Western blot法对重组腺病毒感染后的细胞上清液进行检测,鉴别插入基因表达产物是否表达。对方法的专属性和耐用性进行验证,并采用该方法鉴别3批重组腺病毒疫苗样品插入基因表达产物的活性。结果重组腺病毒疫苗插入基因表达产物能被黏蛋白1-生存素[mucin 1(MUC1)-Survivin]的两种抗体分别识别,在相对分子质量约37000处可见特异性条带,大小和组成与实际值相符,方法的专属性和耐用性均符合要求。3批重组腺病毒疫苗插入基因表达产物也均能被MUC1-Survivin的两种抗体分别识别,在相对分子质量约37000处可见特异性条带,而阴性对照不能被识别。结论采用Western blot法检测插入基因表达产物的活性,专属性和耐用性均较好,适用于重组腺病毒疫苗产品的鉴别试验,可为疫苗制备过程中的质量控制及产品检定提供参考。Objective To develop and validate a method for identification of expressed products of inserted gene in recombinant adenovirus vaccine.Methods The cell supernatant infected with recombinant adenovirus was determind by Western blot to identify the expression of inserted gene.The method was verified for specificity and durability,by which the activity of expressed products of inserted genes in three batches of recombinant adenovirus vaccine samples were identified.Results The expressed product of inserted gene in recombinant adenovirus vaccine was recognized by two antibodies of mucin 1(MUC1)-Survivin.Specific bands each with a relative molecular mass of about 37000 were observed,of which the size and composition were consistent with the actual values,and the specificity and durability of the method met the relevant requirements.The expressed products of inserted gene in three batches of recombinant adenovirus vaccine samples were also recognized by both the two antibodies,which showed specific bands each with a relative molecular mass of about 37000,while the negative control was not recognized.Conclusion Western blot shows good specificity and durability in determination of the activity of expressed products of inserted gene,which is suitable for the identity test of recombinant adenovirus vaccine products.It provides a reference for the quality control of product in the process of vaccine preparation.

关 键 词：腺病毒疫苗 基因表达 鉴别试验 

分 类 号：Q789[生物学—分子生物学]