| 引用本文: | 李剑波,潘海桦,姜云水,吴洁,陈刚,高孟,金素凤,吴小红,包佳源,陈科达,庄昉成.人乳头瘤病毒重组蛋白疫苗酶联免疫检测法的建立及其应用[J].中国现代应用药学,2015,32(1):10-14. |
| LI Jianbo,PAN Haihua,JIANG Yunshui,WU Jie,CHEN Gang,GAO Meng,JIN Sufeng,WU Xiaohong,BAO Jiayuan,CHEN Keda,ZHUANG Fangcheng.Establishment and Application of ELISA Method for HPV16 L2E6E7 Vaccine[J].Chin J Mod Appl Pharm(中国现代应用药学),2015,32(1):10-14. |
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| 人乳头瘤病毒重组蛋白疫苗酶联免疫检测法的建立及其应用 |
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李剑波1, 潘海桦2, 姜云水1, 吴洁1, 陈刚1, 高孟1, 金素凤2, 吴小红1, 包佳源2, 陈科达1, 庄昉成1
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1.浙江省医学科学院病毒病研究所，杭州 310013;2.浙江普康生物技术有限公司，杭州 310013
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| 摘要: |
| 目的 建立人乳头瘤病毒重组蛋白疫苗(HPV16 L2E6E7)酶联免疫的检测方法，用于疫苗发酵过程中的重组蛋白表达的监控。方法 用常规方法制备兔抗HPV16 L2E6E7多克隆抗体作为包被抗体，商品化小鼠抗HPV16 E7单克隆抗体为检测抗体，夹心ELISA方法检测HPV16 L2E6E7抗原参考品，建立抗原标准曲线，确定线性范围及检测限，同时验证该方法的特异性。结果 建立了检测HPV16 L2E6E7抗原含量的夹心ELISA方法，抗原参考品系列浓度在19.53~1 250 ng·mL-1内有很好的线性(R2>0.99)和回收率(90%~110%)；特异性好，不受发酵液中宿主菌蛋白的干扰。结论 建立了人乳头瘤病毒重组蛋白疫苗重组抗原含量的测定方法，为该疫苗的发酵工艺的质控提供了有效技术手段。 |
| 关键词: 人乳头瘤病毒16型 酶联免疫吸附检测 重组蛋白 抗体 |
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| 基金项目:浙江省医学重点学科群建设(XKQ-010-001)；杭州市科技局重大科技创新项目(20112313A37) |
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| Establishment and Application of ELISA Method for HPV16 L2E6E7 Vaccine |
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LI Jianbo1, PAN Haihua2, JIANG Yunshui1, WU Jie1, CHEN Gang1, GAO Meng1, JIN Sufeng2, WU Xiaohong1, BAO Jiayuan2, CHEN Keda1, ZHUANG Fangcheng1
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1.Institute of Viral Diseases, Zhejiang Academy of Medical Sciences, Hangzhou 310013, China;2.Zhejiang Pukang Biotechnology Co., Ltd., Hangzhou 310013, China
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| Abstract: |
| OBJECTIVE To establish a quantitative assay method for determining the titer of HPV16 L2E6E7 vaccine during its development and manufacturing. METHODS By using a conventional method, rabbit anti-HPV16 L2E6E7 polyclonal antibodies were generated and used together with a commercially available mouse anti-HPV16 E7 monoclonal antibody to establish a sandwich ELISA method for HPV16 L2E6E7 vaccine. The linearity, sensitivity, and specificity of the method were evaluated. RESULTS A specific and sensitive ELISA method for assaying HPV16 L2E6E7 vaccine quantitatively was developed. Good linearity was shown within the detection range of 19.53-1 250 ng·mL-1(R2>0.99), and the recovery rate was 90%-110%. The method showed strong sensitivity and high specificity with no interference by host cell proteins of E. coli. CONCLUSION A quantitative ELISA method is established for HPV16 L2E6E7 vaccine. It offers a useful technical tool for quality control of the vaccine during its fermentation process. |
| Key words: human papillomavirus 16 (HPV16) ELISA recombinant protein antibody |
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