鸡抗内毒素卵黄抗体IgY的制备

Preparation and screening of anti-endotoxin IgY in vitro

目的 研究分别应用内毒素 (LPS)、类脂A(LipidA)和大肠杆菌突变株J5免疫鸡后其蛋黄中抗内毒素抗体IgY的产量、纯度、效价并筛选最佳免疫抗原。方法 分别应用内毒素 (LPS)、类脂A(LipidA)和大肠杆菌J5突变株作为抗原免疫 2 5周龄Leghorn鸡 ,水溶法 (WD)提取蛋黄中抗体IgY ,双紫外光测定抗体含量 ,SDS PSGE电泳检测抗体纯度 ,细菌酶联染色和ELISA检测抗体特异性、效价及筛选最佳免疫抗原。结果 3种抗内毒素IgY含量和效价分别为 14 4mg ml和 1∶12 80 0 (J5 )、10 61mg ml和 1∶12 80 0(LPS)、9 2 6mg ml和 1∶3 2 0 0 (LipidA) ,抗体纯度均为 95 %左右。结论 大肠杆菌突变株J5和内毒素 (LPS)为最佳免疫抗原 ,免疫鸡后其蛋黄中抗内毒素抗体IgY的产量和效价最高。

Objective To observe yield, purity and titer of anti-endotoxin IgY isolated from 3 kinds of eggs yolk of hens immunized respectively with J5 mutant of E. coli , lipopolysaccharide (LPS) and lipid A. Methods Laying hens (White Leghorn), 25 weeks old, were immunized respectively with J5 mutant of E.coli, lipopolysaccharide and lipid A. Three kinds of IgY were purified respectively by using the water dilution method (WD), and their purity and yield were measured respectively with SDS PAGE and ultraviolet spectrophotometry. The titer of IgY and specificity of IgY was assayed with ELISA and enzyme immunochemistry respectively. Results Yield and ELISA titer of 3 kinds of IgY were 14.4 mg/ml and 1∶12 800 for J5, 10.61 mg/ml and 1∶12 800 for LPS, 9.26 mg/ml and 1∶3 200 for Lipid A respectively. The purities of the IgY were all about 95%. Conclusion The yield and titer of anti endotoxin IgY from eggs yolk of laying hens immunized respectively with J5 mutant of E. coli and LPS are much more higher than those from Lipid A group. J5 mutant and LPS are two kinds of better antigen to stimulate anti endotoxin antibody IgY.