CAJ | 학술논문

用猪流行性腹泻病毒（PEDV）免疫蛋鸡后提取抗猪流行性腹泻病毒的特异性卵黄抗体为捕获抗体，鼠抗猪流行性腹泻病毒多克隆抗体为检测抗体，建立了猪流行性腹泻病毒病原检测的双抗夹心ELISA，其最适包被抗体浓度为1：4000（12.35μg/mL），鼠抗猪流行性腹泻病毒的多克隆抗体最佳浓度为1：6000（10.25μg/mL），样品反应时间为30min，酶标抗体工作浓度为1：6000，并以OD450≥0.130作为阳性判断标准。该方法与猪传染性胃肠炎、猪轮状病毒、猪流感病毒、大肠杆菌K88、K99、987P、F41等病原无交叉反应。对经猪流行性腹泻病毒PCR检测的100份粪便样本进行检测表明，8份PCR检测阳性样本中6份为本法阳性；PCR阴性者本法全部阴性。实验结果表明该方法具有良好的特异性和敏感性，可用于猪流行性腹泻病毒的快速检测。
용저류행성복사병독（PEDV）면역단계후제취항저류행성복사병독적특이성란황항체위포획항체，서항저류행성복사병독다극륭항체위검측항체，건립료저류행성복사병독병원검측적쌍항협심ELISA，기최괄포피항체농도위1：4000（12.35μg/mL），서항저류행성복사병독적다극륭항체최가농도위1：6000（10.25μg/mL），양품반응시간위30min，매표항체공작농도위1：6000，병이OD450≥0.130작위양성판단표준。해방법여저전염성위장염、저륜상병독、저류감병독、대장간균K88、K99、987P、F41등병원무교차반응。대경저류행성복사병독PCR검측적100빈분편양본진행검측표명，8빈PCR검측양성양본중6빈위본법양성；PCR음성자본법전부음성。실험결과표명해방법구유량호적특이성화민감성，가용우저류행성복사병독적쾌속검측。
Porcine epidemic diarrhea virus(PEDV)was used to immunize laying hens for preparation of speciifc yolk antibody(IgY)as capturing antibody. A double antibody sandwich ELISA(DAS-ELISA)was developed for the detection of PEDV in pigs. The optimal coating concentration of anti-PEDV IgY was 1:4000(12.35μg/mL);the opti-mal concentration of mouse-anti-PEDV polyclonal antibody was 1:6000(10.25μg/mL);optimal reaction time was 30 minutes;optimal working concentration of HRP-labelled goat-anti-mouse IgG was 1:6000;and the positive standard value was OD450≥0.130. The DAS-ELISA showed no cross-reaction with porcine transmissible gastroenteritis virus, porcine rotavirus,Escherichia coli k88/k99/987P/F41. One hundred clinical fecal samples which had been detected by real-time PCR were analyzed by this method. 6 out of 8 real-time PCR positive samples were positive in the DAS-ELISA,and all the negative samples in real-time PCR were negative in the DAS-ELISA. The results indicated that the DAS-ELISA was highly speciifc,reproducible and sensitive,and suitable for rapid detection of PEDV.