摘要
为建立快速检测禽类黏膜组织中SIgA含量的方法,本研究原核重组表达鸡SIgA重链片段蛋白,纯化后免疫BALB/c鼠。以鸡胆汁中分离纯化的SIgA作为检测抗原,常规单克隆技术筛选出1株能够稳定分泌抗鸡SIgA单克隆抗体(MAb)的IgG2a、κ型MAb。经ELISA和western blot分析,所获得的1株MAb亲和力高、特异性强;采用生物素标记纯化MAb腹水IgG为检测二抗,以禽流感-新城疫重组二联活载体疫苗免疫SPF鸡为模型,初步建立了新城疫病毒和H5亚型禽流感病毒特异的黏膜SIgA间接ELISA检测方法。本研究为开展特异性家禽黏膜免疫机制的研究提供了重要技术手段。
To establish the method for detection of specific mucosal SlgA, BALB/c mice was immunized with puriticdrecombinant chicken partial SIgA heavy protein expressed in E. coli. A hybridoma cell line which steadily secreted monoclonalantibody （MAb） against chicken SIgA was prepared by fusion the mouse spleen cells with SP2/0 cells and screened by H.ISA coating with SIgA from chicken bile. Isotope of the MAb was lgG2a, K type. ELISA and western blot showed the MAb was highaffinity and specificity. An indirect ELISA method based on biotin-labeled MAb for the detection of Newcastle disease virus （NDV） and H5 avian influenza virus （H5 AIV） specific mueosal SIgA was established, and the method was successfully used forthe detection of NDV and H5 AIV specific SIgA in salivary of chickens immunized with recombinant NDV live vaccine. Our study provided technological evidence for mucosal immunity mechanism in poultry.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2012年第2期138-142,共5页
Chinese Journal of Preventive Veterinary Medicine
基金
国家攻关项目(2004BA519A19
2005BA711A10)
国家973项目(2005CB523200)
