抗迟钝爱德华氏菌卵黄抗体制备及其抑菌、吞噬调理活性分析  

Preparation of IgY and its antibacterial,phagocytize opsonize effects against Edwardsiella tarda

作　　者：顾叶华 李登峰[1] 周永强[1] GU Ye-hua;LI Deng-feng;ZHOU Yong-qiang(Applied Marine Biotechnology Key Laboratory of the Ministry of Education,Ningbo University,Ningbo 315211,China)

机构地区：[1]宁波大学应用海洋生物技术教育部重点实验室,浙江宁波315211

出　　处：《宁波大学学报（理工版）》2018年第5期1-6,共6页Journal of Ningbo University：Natural Science and Engineering Edition

基　　金：国家海洋局开放基金(HY201602);浙江省公益性技术应用研究计划项目(2011C32004);宁波市农业攻关项目(2011C10004);宁波大学王宽诚幸福基金

摘　　要：迟钝爱德华氏菌(Edwardsiella tarda)是常见人畜共患病病原.为获得高效抗E.tarda卵黄抗体(Immunoglobulin of Yolk,IgY),将纯培养的迟钝爱德华氏菌灭活,免疫蛋鸡,由蛋黄纯化IgY,用间接ELISA法检测IgY效价.对制备的IgY进行了抑菌活性检测和动物保护实验:分别将抗E.tarda IgY、普通鸡蛋IgY与E.tarda(2×10~3 CFU·mL^(-1))孵育,涂平板,菌落计数;分别将抗E.tarda IgY、普通鸡蛋IgY与E.tarda(2×108 CFU·mL^(-1))等体积混合后注射克氏原螯虾(Procambarus clarkia),记录各组成活率.检测抗E.tarda IgY的吞噬调理活性:分别将抗E.tarda IgY、普通鸡蛋IgY与E.tarda(103 CFU·mL^(-1))及小鼠血细胞混合后,接种平板培养、计数.结果发现:制备的IgY质量浓度为8.8 mg·mL^(-1),每个鸡蛋可以获得IgY约130 mg,效价为1:10 240;体外相对抑菌率为86.6%;对克氏原螯虾的平均保护率为66.7%;能促进细胞吞噬E.tarda,相对吞噬率为79.8%.Edwardsiella tarda is responsible for causing bacterial disease in variety of animals（reptiles,amphibians,and fishes,including catfish,turbot and eels）.To prepare specific egg yolk antibody（IgY） against it,laying hens were immunized with purified and inactivated E.tarda for three times.IgY were separated and purified from the yolk of the eggs collected one month after injection by acetic acid buffer dilution,salting-out with ammonium sulfate,and dialysis with PBS.Titer of the IgY were detected by indirect enzyme-linked immunosorbent assay.The concentration and the purity of the IgY were detected by OD value and SDS-PAGE.Results show that concentration of purified IgY is 8.8 mg·mL-1,the mean production from each egg is 130 mg and the titer is 1：10 240.Phagocytize opsonize effects,bacteriostasis in vivo in vitro of the prepared IgY were measured.Rate of bacteriostasis in vitro experiment is 86.6%;The IgY can rise the phagocytize opsonize effect rate to 79.8%;IgY can protect artificially infected crayfish Procambarus clarkia and reduce mortality rate from 100% to 66.7%.

关 键 词：迟钝爱德华氏菌 卵黄抗体 制备 抑菌活性 吞噬调理 

分 类 号：S948[农业科学—水产养殖]