邻苯二甲醛(OPA)法检测铝吸附疫苗中抗原含量

中国生物工程杂志

2011, Vol. 31

Issue (7): 109-113

技术与方法

邻苯二甲醛(OPA)法检测铝吸附疫苗中抗原含量

雷宁¹, 张远², 肖海波², 曾庆乐¹

1. 成都理工大学材料与化学化工学院成都 610059;
2. 成都海圻生物科技有限公司成都 610041

Determination of Protein Content of Alhydrogel-absorbed Vaccine Using O-phthalaldehyde (OPA) Fluorometric Method

LEI Ning¹, ZHANG Yuan², XIAO Hai-bo², ZENG Qing-le¹

1. College of Material and Chemistry & Chemical Engineering, Chengdu University of Technology, Chengdu 610059, China;
2. Chengdu Fronter BioScience Inc., Chengdu 610041, China

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摘要：

目的:建立快速测定铝胶吸附的疫苗中抗原含量的OPA荧光检测法。方法:利用邻苯二甲醛(OPA)在2-巯基乙醇存在下与氨基酸的N端或L-谷氨酸侧链反应,在460nm处生成荧光衍生物的原理,建立了无需进行抗原蛋白提取的检测方法,并对该方法的特异性、线性、精密度、回收率、重复性进行考察。同时,配合钠十二烷基的硫酸盐-聚丙烯酰胺凝胶电泳法(SDS-PAGE),研究在配制过程中抗原蛋白与佐剂是否分离。结果: OPA荧光法,在抗原蛋白含量为0.02~0.16mg/ml时,线性良好,准确度达90%~115%,批内和批间相对标准偏差≤15%。电泳结果显示,CpG的加入可能会导致少量抗原蛋白从铝胶中解离。结论:该方法快速、准确、灵敏度较高,重复性好,可以应用于含铝胶疫苗制剂的实验室甚至生产质控过程。

关键词： 邻苯二甲醛; 铝胶; 疫苗

Abstract:

Objective: To develop a generic method for the determination of protein content in alhydrogel (aluminum) absorbed vaccine formulation using o-phthalaldehyde (OPA). Method: based on the theory that OPA can react with amino acids under alkaline conditions and give strong fluorescence, in the presence of 2-mercaptoethanol, a fluorescent method without antigen extraction was developed and the specificity, linearity range, accuracy and precision were evaluated. The integrity of the vaccine formulation was also investigated by SDS-PAGE. Results: a good linearity was obtained from 0.02mg/ml to 0.16mg/ml of antigen content, the accuracy of 90%~115% and CV% less than 15% were achieved. Little protein disassociation was also observed after CpG was added into the formulation. Conclusion: the OPA fluorescent method was simple and can be further used not only in the laboratory but also in routine quality control of the vaccine formulation with alhydrogel.

Key words: O-phthalaldehyde (OPA) Alhydrogel Vaccines

收稿日期: 2010-12-21 出版日期: 2011-07-25

ZTFLH:

Q819

基金资助:

国家自然科学基金(30940065)、山东省自然科学基金(Y2008C06)资助项目

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引用本文:

雷宁, 张远, 肖海波, 曾庆乐. 邻苯二甲醛(OPA)法检测铝吸附疫苗中抗原含量[J]. 中国生物工程杂志, 2011, 31(7): 109-113.

LEI Ning, ZHANG Yuan, XIAO Hai-bo, ZENG Qing-le. Determination of Protein Content of Alhydrogel-absorbed Vaccine Using O-phthalaldehyde (OPA) Fluorometric Method. China Biotechnology, 2011, 31(7): 109-113.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2011/V31/I7/109

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