新城疫病毒单克隆抗体研究Ⅲ酶标单克隆抗体ELISA夹心法检测鸡新城疫病毒抗原

The Research and Application of the Monoclonal Antibody of Newcastle Disease Virus(NDV)Ⅲ.The Determination of the Antigens of Chick NDV by the EnzymeLabeled Mono clonal Antibody Elisa Sandwich Method

应用双抗体ＥＬＩＳＡ夹心法检测禽类新城疫病毒（ＮＤＶ）抗原，用单克隆抗体或多克隆抗体包被固相载体，将活化的４０孔板作为捕捉待检新城疫病毒抗原，以辣根过氧化物酶（ＨＲＰ）与单克隆抗体的结合物作为示踪抗体，比较了单克隆抗体和多克隆抗体的捕捉效率。结果表明，ＮＤＶ单抗２Ｃ１２、４Ｈ１０优于血清ＩｇＧ，同时确定了双抗体ＥＬＩＳＡ夹心法对不同日龄健康鸡组织产生背景反应的Ｏ．Ｄ．值基本参数，以Ｘ＋２ＳＤ为判定阈值。捕捉抗体２Ｃ１２、４Ｈ１０的工作浓度为５００～４００μｇ／ｍｌ；示踪抗体２Ｃ１２、４Ｈ１０的工作浓度为１∶３０００～１∶２０００，ＥＬＩＳＡ夹心法的灵敏度可达４μｇＮＤＶ蛋白／ｍｌ；对ＮＤＶ不同毒株感染鸡各脏器的ＮＤＶ检出率为：肾１００％、肺９０％。证明用双抗体ＥＬＩＳＡ夹心法检测鸡新城疫病毒抗原特异，敏感。

The research determine the antigens of avian Newcastle disease virus(NDV) with Double Antibody ELISA Sandwich Method and wrap up solid carriers by monoclonal or polyclonal antibodies and seize antigens of ndv in activated 40hole plates and use conjugates of Horseradish Peroxidase(HRP) and monoclonal antibodies as tracer antibodies.The seizing efficiencies between the monoclonal and polyclonal antibodies are compared,which show that NDV monoclonal antibodies 2C 12 ,4H 10 are superior to polyclonal I gG ;The basic parameter of O.D. producing background reaction in tissues of different day's old healthy chicks by Double Antibody Sandwich ELISA Method is determined,i.e. +2SD. The working concentrations of the seizing antibodies 2C 12 ,4H 10 and the tracer antibodies 2C 12 ,4H 10 are 500～400 μg/ml and 1∶3 000～1∶2 000 ,respectively.The sensitivity of Sandwich ELISA may attain 4 μg/ml NDV protein.The examination rates of NDV in various tissues of the chicks infected by different strains of NDV are 100% and 90% in kidneys and lungs,respectively,the research confirm that Double Antibody ELISA Sandwich Method is distinctive and sensitive in the determination of the antigens of NDV,which has supplied a new way for the examination of NDV.