摘要
目的:制备特异性抗肠道病毒71型(Enterovirus 71,EV71)的卵黄抗体并检测其生物学性能,这对EV71感染手足口病的预防和治疗具有重大的意义。方法:用纯化并灭活后EV71作为抗原免疫健康产蛋鸡。采用本实验室水提综合聚乙二醇法提取纯化鸡卵黄抗体;用Bradford法测定卵黄抗体提取液的蛋白含量;还原性SDS-PAGE凝胶电泳分析测定提取蛋白的纯度及相对分子质量;分别用ELISA、双向免疫琼脂扩散检测纯化特异性卵黄抗体抗EV71效价;Western blot分析特异性卵黄抗体的免疫反应性。结果:卵黄中卵黄抗体含量达7.76 mg/ml,卵黄抗体的纯度为94.86%。初免10天后蛋黄中可检测到特异性抗EV71卵黄抗体,初免40天后ELISA检测抗体效价高达1∶20 480,双向琼脂扩散检测效价达1∶16。提取的卵黄抗体具有良好的免疫反应性,能与EV71特异性结合。结论:水提综合聚乙二醇法提取纯化得到的抗EV71卵黄抗体产量和纯度高、效价好且具有良好的免疫反应性。
Objective：Preparing specificity chicken egg yolk antibody（IgY） against enterovirus 71（EV71） and detecting its biological activities,this is significant for prevention and treatment of EV71 infection.Methods：Four hens were immunized three times with the purified human EV71 virus as an antigen.The immunoglobulin Y（IgY） antibody was extracted and purified from the egg yolk by water combine polyethylene glycol（water-peg） method.The yield and molecular mass of IgY were analysed by Bradford method and SDS-PAGE electrophoresis.ELISA and Agar double immunodiffusion were used for detect the titer of IgY,while the immunoreactivity of anti-EV71 IgY was tested using Western blot.Results：The yield and purity of IgY were 7.76 mg/ml and 94.86% respectively.Results of ELISA indicated that the specific anti-EV71 IgY antibody appeared in the egg yolk 10 days after primary immunization and reached the highest titer 1∶20 480 after 40 days,while the titer of Agar double immunodiffusion was 1∶16.Western blot analysis revealed that the purified IgY antibody owned good immunoreactivity and a specifically combination capacity of EV71 virus.Conclusion：Egg yolk is a large scale source of antidodys specifically against EV71 virus.The anti-EV71 IgY extracted and purified by Water-PEG method possess good production,purity,and biological activities.
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2011年第10期913-916,921,共5页
Chinese Journal of Immunology
基金
广东省211工程计划
天河区科技计划(No.104G193)资助
关键词
肠道病毒71型
卵黄抗体
手足口病
制备
鉴定
Enterovirus 71
IgY
Hand-foot-and-mouth disease
Preparation
Identification
