鸡白细胞介素18双抗体夹心ELISA检测方法的建立及初步应用

应用识别不同表位的鸡白细胞介素18成熟蛋白（Mature chicken interleukin-18，mChIL-18）的2株单克隆抗体（mAb）1G9和2E6，建立检测mChlL-18的双抗体夹心ELISA，并利用此方法对禽网状内皮组织增生症病毒（Reticuloendotheliosis virus，REV）人工感染SPF鸡体内mChIL-18的分泌水平进行检测。结果显示，捕获抗体的最佳质量浓度为8mg／L，检测抗体的工作效价为1：800，待检样品的最佳稀释度为1：400，检测敏感度可达31．5ng／L，与其他细胞因子等抗原蛋白无交叉反应；跟对照组相比，REV感染鸡体内mChIL-18的表达量在7、14、21、28、35、42、49d均呈现升高，但只有14日龄时表现差异显著（P〈0．05）。结果表明，本试验成功建立了ChIL-18的双抗体夹心ELISA，为鸡传染病的细胞免疫学研究提供了可靠方法。

Development and preliminary application of double sandwich ELISA for detecting chicken interleukin-18

Double sandwich ELISA for mature chicken interleukin 18（mChIL-18） was developed using mAb 2E6 as a capture antibody and HRP-labeled mAb 1G9 as a detection antibody and then the expression levels of mChIL-18 in serum samples of SPF chickens infected with reticuloendo- theliosis virus（REV）were investigated by using this method. The results were as follows.the optimal concentration of capture antibody was 8 rag/L, the working concentration of detection antibody was 1 ： 800, the best dilution of the samples was 1 ： 400, and the detection limit of the sandwich ELISA for mChIL-18 was 31.5 ng/L, there was no cross reaction with other antigen protein including chicken cytokines; mChIL-18 expression levels were higher in the serum samples of SPF chickens from treatment group than those in the control group at 7,14,21,28,35,42 and 49 d, but only the samples of 14 d were significantly changed（P〈0.05）. This method can be useful to understand the role of this cytokine in various chicken diseases and develop specific cell- mediated immunityassay.