英文文献汇报_精品文档.ppt

1、Identification of a linear B-cell epitope on the avian leukosis virus P27 protein using monoclonal antibodies,Xiaofei Li 1,2,Liting Qin 2,Haibo Zhu 3,Yingjun Sun 2,Xuezhi Cui 2,Yadong Gao 2,Xiaole Qi 1,Yongqiang Wang 1,Honglei Gao 1,Yulong Gao 1,Xiaomei Wang 1Arch VirolIF=2.058,Abstract Avian leukos

2、is virus(ALV)is an avian oncogenic retrovirus that can induce various clinical tumors.The capsid protein P27 is the group-specific antigen of ALV and has many viral antigen sites that are easy to detect.In this study,we produced a monoclonal antibody(mAb),3A9,that is specific for the P27 protein.A s

3、eries of partially overlapping peptides were screened to define 181PPSAR185 as the minimal linear epitope recognized by mAb 3A9.The identified epitope could be recognized by chicken anti-ALV and mouse anti-ALV P27 sera.The epitope was highly conserved among a number of ALV-A,ALV-B and ALV-J strains.

4、MAb 3A9 might be a valuable tool for the development of new immunodiagnostic approaches for ALV,and the defined linear epitope might help further our understanding of the antigenic structure of the P27 protein.,Avian leukosis viruses(ALVs),which belong to the family Retroviridae,are classified into

5、A,B,C,D,E,and J subgroups1,2.The A,B and J subgroups of ALV are the most common ALVs in commercial poultry,whereas subgroups C and D have rarely been reported.Subgroup E is a ubiquitous endogenous leukosis virus of low pathogenicity 3.ALV predominantly causes lymphocytic leukemia,myeloid leukosis an

6、d other sarcomas and can also lead to immunosuppression effects,such as the abnormal development of immune organs,growth retardation,and decreased immune responses.4,5.ALV subgroup J(ALV-J)was first isolated in the UK from white meat-type chickens in 1988 2.In China,ALV-J was first reported in 1999.

7、Since then,ALV-J isolates have been detected in layer chickens,broiler breeders and local chickens throughout most regions of China and have caused various tumors in chickens 68.In recent years,ALV-J has become widespread and has resulted in severe economic losses to the poultry industry 6,8.,1.Intr

8、oduction,Effective vaccines against ALV are not available.The control of ALV infection occurs primarily by establishing exogenous ALV-free poultry flocks by adopting eradication as the strategy of choice 9.Because of substantial antigenic and genetic variation among ALV isolates and high levels of v

9、ertical and horizontal transmission,eradication has been difficult 10,11.Thus,effective methods for the accurate detection of ALV antigens in chickens are critical for the control of ALV infections.The genome of ALV consists of 5-LTR-UTR-gag-polenv-UTR-LTR-3.The gag,pol and env genes are the viral s

10、tructural genes 11.The P27 protein,which is encoded by the gag gene,is the capsid protein and acts as a group-specific antigen.The ALV P27 gene is highly conserved and exhibits 96%sequence identity among the exogenous subgroups(A,B,C,D and J).In addition,the P27 protein content accounts for more tha

11、n 30%of the viral protein,and P27 has many viral antigen sites that are easy to detect.Therefore,the P27 protein is the first choice for preparing antibodies for detection.Epitopes of proteins are classified as either continuous or discontinuous depending on whether the amino acids included in the e

12、pitope are continuous in the peptide chain or not1214.Continuous epitopes sometimes do not represent the entire antigenic epitope in the viral protein but instead only a short cross-reactive portion of a larger,discontinuous epitope 12,13.,B-cell epitopes are regions that are recognized by the bindi

13、ng sites or paratopes of antibody molecules when they are present in their free form in serum or as membrane-bound B-cell receptors 12.Correct identification of B-cell epitopes within an antigenic protein may open the door for the design of molecules that mimic potentially protective epitopes and co

14、uld be used to raise specific Abs or be used as prophylactic or therapeutic vaccines 1517.Identification of B-cell epitopes could promote protective immunity in the context of emerging and re-emerging infectious diseases and potential bioterrorist threats 15.The aim of this study was to generate a P

15、27-specific antibody against ALV using purified recombinant P27 protein as the immunogen and subsequently to identify the B-cell epitope recognized by the antibody.The information described in this study will facilitate the development of ALV diagnostic tools and will further our understanding of th

16、e antigenic structure of the P27 protein.,2.Materials and methods,重组质粒的构建 构建重叠片段短肽,免疫小鼠制备单抗,IFA&Western blot检测单抗特异性,鉴定最小抗原表位基序,分析表位保守性,3.Results,Fig.1 Characterization of the recombinant P27 protein and mAb 3A9.(A)Expression and purification of the recombinant P27 protein.M,molecular marker(kDa);1,recombinant P27 protein before IPTG induction;2,recombinant P27 protein after IPTG induction;3,purified recombinant P27 protein.(B)Recognition of the purified ALV-J P27 protein by mAb 3A9 in western bl