副溶血性弧菌高免卵黄抗体的制备和不同提纯方法效果的比较

doi:10.13481/j.1671-587x.20170245

吉林大学学报(医学版) ›› 2017, Vol. 43 ›› Issue (02): 441-445.doi: 10.13481/j.1671-587x.20170245

副溶血性弧菌高免卵黄抗体的制备和不同提纯方法效果的比较

翟玥, 曲笑锋, 庞博, 李丽, 徐坤, 王梦涵, 高孜博, 宋佳音, 宋秀玲, 金明华

吉林大学公共卫生学院卫生检验学教研室, 吉林长春 130021

收稿日期:2016-09-13 出版日期:2017-03-28 发布日期:2017-03-31
通讯作者: 宋秀玲,讲师(Tel:0431-85619441,E-mail:songxiuling@jlu.edu.cn);金明华,副教授,硕士研究生导师(Tel:0431-85619441,E-mail:jinmh@jlu.edu.cn) E-mail:songxiuling@jlu.edu.cn;jinmh@jlu.edu.cn
作者简介:翟玥(1992-),女,吉林省长春市人,在读医学硕士,主要从事卫生检验技术方面的研究。
基金资助:
国家自然科学基金青年基金资助课题（81502849，81401721）；吉林大学研究生创新基金资助课题（2016040）；吉林大学大学生创新创业训练计划项目资助课题（2015721031）

Preparation of high immunity yolk antibody against Vibrio parahemolyticus and comparison of effectiveness between different extraction methods

ZHAI Yue, QU Xiaofeng, PANG Bo, LI Li, XU Kun, WANG Menghan, GAO Zibo, SONG Jiayin, SONG Xiuling, JIN Minghua

Department of Health Inspection, School of Public Health, Jilin University, Changchun 130021, China

Received:2016-09-13 Online:2017-03-28 Published:2017-03-31

摘要/Abstract

摘要： 目的：通过比较聚乙二醇（PEG）沉淀法、氯仿抽提法和氯仿/PEG法3种卵黄抗体（IgY）提纯方法的提纯效果，为批量提纯高质量IgY提供依据。方法：制备副溶血性弧菌灭活疫苗，采用肌肉多点注射法免疫高产蛋鸡，收集鸡蛋，分别采用PEG沉淀法、氯仿抽提法和氯仿/PEG法对鸡蛋中的IgY进行提纯，比较3种提纯方法获得的IgY的蛋白质量浓度、抗体效价和纯度，结合操作过程、成本效益及安全性对3种提纯方法进行综合分析比较。结果：3种方法所提纯的IgY中，蛋白质量浓度从高到低依次为氯仿抽提法、氯仿/PEG法、PEG法；抗体效价相差不大，氯仿抽提法稍高；抗体纯度由高到低依次为PEG法、氯仿/PEG法、氯仿抽提法。PEG法安全性较好，但提取效率相对较低，成本投入较高；氯仿抽提法能够高效地从卵黄中获得IgY，但提取物中掺杂了大量的杂质蛋白；氯仿/PEG法提纯效率高，抗体纯度好，成本效益相对较高。结论：PEG法适用于实验室中少量IgY的提取；氯仿/PEG法的提取效率高，抗体纯度好，可用于批量提取高质量的IgY。

关键词: 副溶血性弧菌, 氯仿法, 鸡卵黄抗体, 聚乙二醇法

Abstract: Objective: To compare the effectiveness between three methods for purifying the immunoglobulin of egg yolk(IgY) which are polyethylene glycol (PEG) method, chloroform extraction method and chloroform/PEG method, and to provide basis for obtaining the batch of IgY. Methods: The inactivated vaccine of Vibrio parahemolyticus (V. parahemolyticus) was prepared and the hens were immunized by multi-point intramuscular injection.The eggs were collected and the IgY was purified by PEG method, chloroform extraction method and chloroform/PEG method.The protein extraction rate, the IgY titer and the purity of the antibody which purified by different methods were detected.Furthermore, the operation process, cost and safety of the three methods were analyzed. Results: The protein contents of the extraction belonging three methods from high to low in turn were chloroform extraction method, chloroform/PEG method, and PEG method.There was no significant difference in the antibody titer between three methods, and the tiler of chloroform extraction method was slightly high.The purities of purified antibody from high to low in turn were PEG method, chloroform/PEG method and chloroform method.The PEG method had better security but relatively lower extraction efficiency and higher cost.The chloroform/PEG method had high extraction efficiency and good antibody purity. Conclusion: The PEG method is suitable for a small amount of extraction in the laboratory.The chloroform/PEG method is appropriate for extracting the high quality IgY in a batch as it has high extraction efficiency and good antibody purity.

Key words: immunoglobulin of egg yolk, polyethylene glycol method, chloroform method, Vibrio parahemolyticus

中图分类号:

S852.43

翟玥, 曲笑锋, 庞博, 李丽, 徐坤, 王梦涵, 高孜博, 宋佳音, 宋秀玲, 金明华. 副溶血性弧菌高免卵黄抗体的制备和不同提纯方法效果的比较[J]. 吉林大学学报(医学版), 2017, 43(02): 441-445.

ZHAI Yue, QU Xiaofeng, PANG Bo, LI Li, XU Kun, WANG Menghan, GAO Zibo, SONG Jiayin, SONG Xiuling, JIN Minghua. Preparation of high immunity yolk antibody against Vibrio parahemolyticus and comparison of effectiveness between different extraction methods[J]. Journal of Jilin University Medicine Edition, 2017, 43(02): 441-445.

参考文献

[1] 闫俊, 赵小玲, 陈伟强, 等.抗日本血吸虫Sjp40鸡卵黄抗体的制备及其生物学特性鉴定[J].解放军预防医学杂志, 2013, 31(6):487-489.
[2] Guo Y, Gao M, Zhang H, et al.Molecular characterization and B cell membrane expression analysis of Fc fragment gene of goose IgY[J].Res Vet Sci, 2014, 97(2):288-291.
[3] Mclaren RD, Prosser CG, Grieve RC, et al.The use of caprylic acid for the extraction of the immunoglobulin fraction from egg yolk of chickens immunised with ovine alpha-lactalbumin[J].J Immunol Methods, 1994, 177(1/2):175-184.
[4] 段琳, 赵文锋, 丁建坤, 等.卵黄抗体开发现状及应用进展[J].中国生化药杂志, 2010, 31(3):212-215.
[5] Duan HL, He QY, Zhou B, et al.Anti-trimeresurus albolabris venom IgY antibodies:preparation, purification and neutralization efficacy[J].J Venom Anim Toxins Incl Trop Dis,2016, 22(1):23.
[6] 蒋兴龙, 江国永, 成国祥.几种从鸡血浆中提取免疫球蛋白IgY方法的比较[J].饲料与畜牧, 2015 (3):64-67.
[7] 张彩虹, 祝贺, 高又文, 等.抗非洲猪瘟病毒VP73蛋白鸡卵黄抗体的制备及鉴定[J].中国兽医科学, 2015, 45(8):821-825.
[8] 高建, 周玉春, 黄宇烽.鸡卵黄抗体的纯化及临床应用[J].中华男科学杂志, 2008, 14 (2):166-170.
[9] Pauly D, Chacana PA, Calzado EG, et al.IgY technology:extraction of chicken antibodies from egg yolk by polyethylene glycol (PEG) precipitation[J].J Vis Exp, 2011(51):3084.
[10] 林婷婷, 王全溪.两种卵黄抗体不同提取方法的比较试验[J].福建畜牧兽医, 2015, 37 (3):3-6.
[11] Polson A.Isolation of IgY from the yolks of eggs by a chloroform polyethylene glycol procedure[J].Immunol Invest, 1990, 19(3):253-258.
[12] Nasiri K, Nassiri MR, Tahmoorespur M, et al.Production and characterization of egg yolk antibody (IgY) against recombinant VP8-S2 antigen[J].Pol J Vet Sci,2016, 19(2):271-279.
[13] Lévesque S, Martinez G, Fairbrother JM.Improvement of adjuvant systems to obtain a cost-effective production of high levels of specific IgY[J].Poult Sci, 2007, 86(4):630-635.
[14] 刘瑞生, 张洪波, 薛掌林, 等.鸡新城疫卵黄抗体提取方法筛选研究[J].国外畜牧学-猪与禽, 2014, 34 (6):57-59.
[15] 侯亚琴, 贾丽云, 王丽.卵黄抗体检测法在禽流感免疫监测中的应用[J].当代畜牧, 2009(5):15-16.
[16] 陈志蓉, 裴新荣, 张凤兰, 等.氯仿毒性的研究进展及法规管理现状[J].癌变·畸变·突变, 2014, 26(1):71-74.
[17] Adaramoye OA, Azeez AF, Ola-Davies OE.Ameliorative effects of chloroform fraction of Cocos nucifera L.husk fiber against cisplatin-induced toxicity in rats[J].Pharmacognosy Res, 2016, 8(2):89-96.

副溶血性弧菌高免卵黄抗体的制备和不同提纯方法效果的比较

Preparation of high immunity yolk antibody against Vibrio parahemolyticus and comparison of effectiveness between different extraction methods

RichHTML

PDF (PC)

赞

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 1

Metrics

本文评价

推荐阅读 0