吉林大学学报(医学版) ›› 2024, Vol. 50 ›› Issue (2): 303-309.doi: 10.13481/j.1671-587X.20240202

• 基础研究 • 上一篇    

抗单纯疱疹病毒1型的IgY制备及其生物学活性检测

苏海涛1,翟玥2,宋秀玲2,徐坤2,3()   

  1. 1.吉林省疾病预防控制中心寄生虫病预防控制所,吉林 长春 130062
    2.吉林大学公共卫生学院卫生检验教研室,吉林 长春 130021
    3.湖南师范大学医学院食品与营养卫生教研室,湖南 长沙 410013
  • 收稿日期:2023-04-27 出版日期:2024-03-28 发布日期:2024-04-28
  • 通讯作者: 徐坤 E-mail:xu_kun2022@163.com
  • 作者简介:苏海涛(1980-),男,吉林省大安市人,主任医师,医学硕士,主要从事传染病预防控制方面的研究。
  • 基金资助:
    吉林省科技厅科技创新基地(平台)建设项目(YDZJ202302CXJD054)

Preparation of IgY against herpes simplex virus-1 and detection of its biological activity

Haitao SU1,Yue ZHAI2,Xiuling SONG2,Kun XU2,3()   

  1. 1.Institute of Parasitic Disease Prevention and Control,Center for Disease Control and Prevention of Jilin Province,Changchun 130062,China
    2.Department of Hygienic Inspection,School of Public Health,Jilin University,Changchun 130021,China
    3.Department of Food and Nutrition Hygiene,School of Medical Sciences,Hunan Normal University,Changsha 410013,China
  • Received:2023-04-27 Online:2024-03-28 Published:2024-04-28
  • Contact: Kun XU E-mail:xu_kun2022@163.com

摘要:

目的 制备抗单纯疱疹病毒1型(HSV-1)的卵黄抗体(IgY),探讨该抗体的生物学活性,阐明其抗HSV-1的能力。 方法 制备HSV-1灭活疫苗,采用鸡胸多点注射法免疫高产蛋鸡,采用聚乙二醇-6000(PEG-6000)法提纯IgY,采用间接ELISA法、十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)技术和蛋白浓度定量试剂盒测定IgY效价、纯度及蛋白水平,采用间接ELISA法检测IgY中和病毒的能力,测定病毒阻断率,以Vero细胞为病毒感染靶细胞,测定不同抗体浓度(0.015 60、0.031 25、0.062 50、0.125 00、0.500 00、和1.000 00 g·L-1)作用后细胞病变效应(CPE),根据CPE程度测定IgY体外抗HSV-1能力。 结果 制备的IgY效价随免疫时间逐渐升高,达到1/1 024 000后保持稳定;IgY轻链重链条带清晰;IgY平均蛋白水平为11.544 g·L-1;IgY对HSV-1的阻断率可高达77.90%;HSV-1致CPE程度随抗体浓度升高而逐渐降低,当IgY浓度达到0.500 00 g·L-1及以上时,25%以下细胞(甚至无细胞)发生病变。 结论 成功制备出抗HSV-1的IgY,该抗体对HSV-1具有明显的中和阻断能力和体外抑制活性,可用于药物及诊断方法的开发。

关键词: 单纯疱疹病毒1型, 卵黄抗体, 中和病毒, 体外抗病毒能力

Abstract:

Objective To prepare the anti-herpes simplex virus-1 (HSV-1) egg yolk antibody (IgY) and discuss its biological activity, and to clarify its anti-HSV-1 capability. Methods The inactivated HSV-1 vaccine was prepared, the high egg-producing hens were immunized by multi-site injection method in the chicken breast, and IgY was purified by polyethylene glycol-6000 (PEG-6000) method;the titer, purity, and protein levels of IgY were detected by indirect ELISA method, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) technology, and protein concentration quantification kits; the ability of IgY to neutralize the virus was detected by indirect ELISA method; the virus blocking rate was detected.The Vero cells were used as the target cells for viral infection. The cytopathic effect (CPE) at different antibody concentrations (0.015 60, 0.031 25, 0.062 50, 0.125 00, 0.250 00, 0.500 00, and 1.000 00 g·L?1) was detected;the in vitro anti-HSV-1 capacity of IgY was detected according to the degrees of CPE. Results The titer of the prepared IgY was gradually increased with the extension of immunization time. The titer of the IgY stabilized at 1/1 024 000; the clear bands for the IgY light and heavy chains were observed; the average protein level of IgY was 11.544 g·L?1; the blocking rate of IgY against HSV-1 could reach up to 77.90%; the degree of HSV-1 induced CPE was decreased with the increasing of the antibody concentrations, and when the concentration of IgY reached 0.500 00 g·L?1 or higher, less than 25% of the cells (even no cells) exhibited cytopathic changes. Conclusion Anti-HSV-1 IgY is successfully prepared, and this antibody has significant neutralizing and blocking capabilities and in vitro inhibitory activity against HSV-1, which can be utilized for the development of drugs and diagnostic methods.

Key words: Herpes simplex virus-1, Egg yolk antibody, Neutralizing virus, Antiviral ability in vitro

中图分类号: 

  • S852.43