抗弓形虫腺苷激酶IgY的制备及理化性质的研究  

Preparation,identification and physiochemical study of egg yolk immunoglobulin(IgY) against Toxoplasmosis gondii adenosine kinase

作　　者：汪六庆[1] 周银娣[2] 徐海红[1] 

机构地区：[1]合肥市第三人民医院检验科,安徽合肥230061 [2]合肥市第一人民医院检验科,安徽合肥230036

出　　处：《热带医学杂志》2016年第8期958-961,共4页Journal of Tropical Medicine

基　　金：国家重大基础研究计划(2010CB530001);安徽医科大学校科研基金(xkj2015056);合肥市第三人民医院院内课题(2013013)

摘　　要：目的制备抗重组刚地弓形虫腺苷激酶（AK）的抗体IgY,对IgY进行特异性鉴定,并用间接ELISA的方法进行滴度、耐热、耐酸、耐碱、耐胃蛋白酶等理化性质的研究。方法利用弓形虫AK蛋白,免疫产蛋母鸡,制备并纯化抗AK的卵黄抗体IgY,SDS-PAGE和Western blot进行特异性分析,间接EILSA进行滴度和理化性质的检测。结果 SDS-PAGE检测IgY分子量与理论值一致;Western blot显示制备的IgY为抗AK的特异性IgY;二喹啉甲酸（BCA）法测浓度为4.25 mg/ml;间接ELISA检测滴度为1∶1×10~5;耐热,温度80℃时作用5 min不降解;耐碱,pH为8.5时,作用120 min不降解;30%岩藻糖可以完全抑制在pH等于3时的迅速降解;37℃胃蛋白酶作用120 min不降解。结论本研究成功制备并鉴定了抗AK的IgY,IgY具有滴度和浓度高、耐热、耐酸、耐碱、耐胃蛋白酶的性质,为下一步的研究提供了理论依据。Objective To prepare, identify and study the physiochemical properties of egg yolk immunoglobulin（Ig Y）antibodies against Adenosine kinase（AK） of Toxoplasmosis gondii. Methods Recombinant AK protein of Toxoplasmosis gondii was used to immunize hens and prepare anti-AK Ig Y. SDS-PAGE and Western blot were used to determine the specificity of IgY, and the titer and physico-chemical property were determined by indirect ELISA. Results The molecular weight of IgY determined by SDS-PAGE was the same as the predicted molecular weight.Indirect ELISA and Western blotting indicated that Ig Y was specific to AK. The concentration was 5 mg / ml measured by bicinchoninic acid（BCA）; the tire was 1∶1×10~5; it was heat-resistant, lasting 5 min non-degrade at 80℃;base-resistant, lasting 120 min non-degradeat p H 8.5; it degraded quickly when p H was 3, but 30% fucose can suppress the degradation at p H 3; it is resistant to pepsin degradation at 37℃. Conclusion This study prepared and identified Ig Y antibody against AK. It has high titer and high concentration, and was resistant to heat, acid, base and pepsin digestion.

关 键 词：弓形虫 腺苷激酶 卵黄抗体 IGY 

分 类 号：R382[医药卫生—医学寄生虫学]