中文摘要：

中国小麦花叶病是引起小麦（Triticum aestivum）严重减产的中国小麦花叶病毒（Chinese wheat mosaic virus,CWMV）造成的。本研究旨在以制备的抗CWMV特异性单克隆抗体（monoclonal antibody,MAb）为核心建立检测CWMV的血清学方法,为该病毒病的诊断和科学防控提供技术支撑。用CWMV提纯病毒免疫小鼠（Mus musculus）,经细胞融合、培养基筛选培养、阳性细胞的检测与单克隆化后,共获得4株能分泌抗CWMV单克隆抗体的杂交瘤细胞株5H5、7C2、9A7和12E5,并注射入BALB/c小鼠（Mus musculus）腹腔制备单抗腹水;间接酶联免疫吸附实验（enzyme linked immunosorbent assay,ELISA）表明,4种单抗腹水效价均达到10-7,抗体类型及亚类均为Ig G1、kappa链。Western blot分析表明,4株单克隆抗体均能与CWMV的外壳蛋白亚基有特异性反应。利用制备的单抗建立检测田间小麦样品中CWMV的抗原包被酶联免疫吸附实验（antigen coating plate enzyme-linked immunosorbent assay,ACP-ELISA）和斑点酶联免疫吸附实验（dot-enzyme-linked immunosorbent assay,dot-ELISA）,结果表明,该两种血清学方法检测感染CWMV小麦植物组织粗提液呈特异性阳性反应,而检测感染小麦黄花叶病毒（Wheat yellow mosaic virus,WYMV）的小麦、感染大麦黄花叶病毒（Barley yellow mosaic virus,Ba YMV）的大麦（Hordeum vulgare）和健康小麦均呈阴性反应,说明建立的两种方法能特异性地检测小麦植物中的CWMV。灵敏度分析表明,以5H5和12E5单抗为核心建立的ACP-ELISA方法检测小麦病叶的灵敏度达到1∶81 920（W/V,g/m L）,而以7C2和9A7单抗为核心建立的ACP-ELISA方法检测小麦病叶灵敏度达到1∶40 960（W/V,g/m L）倍稀释;以12E5和5H5单抗为核心建立的dot-ELISA方法检测小麦病叶的灵敏度达到1∶2 560倍稀释（W/V,g/m L）,而以7C2和9A7单抗为核心建立的dot-ELISA方法的检测灵敏度达到1∶1 280倍稀释（W/V,g/m L）。田间样品检测?

英文摘要：

Chinese wheat mosaic disease, which is caused by Chinese wheat mosaic virus （CWMV）, leads to severe yield and huge economic losses on wheat （Triticum aestivum）. The purpose of this study was to develop serological methods for detecting CWMV using prepared monoclonal antibodies （MAbs） against CWMV, and to provide technical support for the diagnosis and scientific prevention and control of CWMV in fields. Using the purified CWMV virus particles as an immunogen, 4 hybridoma lines （5H5, 7C2, 9A7 and 12E5） secreting MAbs against CWMV were obtained via cell fusion, cell culture, antibody detection and cell cloning, and injected intraperitoneally into BALB/c mice （Mus musculus） to produce ascitic fluids. The titers of all 4 MAbs in ascites determined by an indirect-ELISA were up to 107. Isotypes and subclasses of all 4 MAbs belonged to IgG1, K light chain. Western blot analysis indicated that all 4 MAbs could specifically react with the coat protein of CWMV. Using the MAbs, antigen-coated plate enzyme-linked immunosorbent assay（ACP-ELISA） and dot-enzyme-linked immunosorbent assay （dot-ELISA） for detecting CWMV were established, respectively. Specificity analysis of the ACP-ELISA and the dot-ELISA indicated that both 2 serological detection methods had positive reactions of detection with CWMV infected wheat plant tissue crude extracts and negative reactions of detection with wheat or barley （Hordeum vulgare） plant infected by Wheat yellow mosaic virus （WYMV） or Barley yellow mosaic virus （BaYMV） and healthy wheat plant. This result suggested that 2 serological detection methods could specifically detect CWMV in wheat plants. Sensitivity analysis showed that the sensitivity of the ACP-ELISA based on the MAbs （5H5 and 12E5） for detecting CWMV in infected wheat tissue could reach 1 ： 81 920（W/V, g/mL）, while the detection sensitivity of ACP-ELISA based on the MAbs （7C2 and 9A7） was up to 1 ： 40 960 （W/V, g/mL）. The sensitivity of the dot-ELISA based on the MAbs ?