传染性法氏囊病病毒VP2基因的表达与单克隆抗体的制备
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摘要
传染性法氏囊病(Infectious Bursal Disease, IBD)是由传染性法氏囊病病毒(Infectious Bursal Disease Virus, IBDV)引起的以侵害雏鸡淋巴组织,特别是中枢免疫器官一法氏囊为主要特征的传染病。该病不仅引起患病动物死亡,还会导致机体免疫抑制,使机体的免疫防御能力降低和疫苗免疫接种失败,每年给养禽业带来巨大经济损失。近年来,由于各地出现的IBDV变异株、强毒株的毒力和抗原性存在一定差异,可逃避现行疫苗的免疫,IBD的流行呈现更为复杂的态势,给该病的防控造成较大困难。IBDV VP2蛋白是病毒主要的结构蛋白和宿主保护性抗原,因此针对VP2蛋白及IBDV检测方法的研究是当前重要的研究内容。本文将流行毒株IBDV VP2基因分别在大肠杆菌和杆状病毒表达系统中进行表达,获得了针对VP2蛋白的单克隆抗体。试验内容主要包括以下三个部分：
1.传染性法氏囊病病毒VP2基因在大肠杆菌中的表达
将从安徽省传染性法氏囊病(IBD)免疫失败的病鸡法氏囊组织中扩增的VP2基因定向克隆入原核表达载体pGEX-4T-1中,构建了原核表达质粒pGEX-VP2,并转化大肠杆菌Rosetta (DE3).经IPTG诱导后,用IBDV双抗体夹心ELISA检测,结果呈阳性反应：SDS-PAGE分析结果显示,重组VP2融合蛋白的分子质量为68 ku,表达量约占菌体总蛋白量的16.0%,表达产物主要以包涵体形式存在于菌体内；Western-blotting结果显示,重组VP2蛋白与IBDV抗体发生反应,形成一条特异性蛋白带,表明获得的重组蛋白具有良好的抗原反应性。
2.传染性法氏囊病病毒VP2基因在昆虫细胞中的表达
为制备针对传染性法氏囊病病毒(IBDV)流行毒株的重组VP2蛋白,将从安徽省IBD免疫失败的病鸡法氏囊组织中扩增的VP2基因插入到杆状病毒表达系统供体质粒pFastBacHTA中,转化大肠杆菌DH10Bac感受态细胞,并成功获得重组杆状病毒表达质粒rBacHT-VP2,用脂质体法转染Sf9细胞,获得重组杆状病毒vBacHT-VP2.经Western-blotting分析,在蛋白分子量约53 ku处出现特异性蛋白条带；以间接免疫荧光试验(IFA)检测vBacHT-VP2感染的Sf9细胞,具有特异性免疫荧光；电镜观察,重组VP2蛋白能够自组装成病毒样颗粒(VLPs)。
3.传染性法氏囊病病毒VP2蛋白单克隆抗体的制备与应用
用大肠杆菌表达的传染性法氏囊病病毒(IBDV)重组VP2蛋白、杆状病毒表达的重组VP2蛋白以及纯化的IBDV分别免疫BALB/c小鼠,利用淋巴细胞杂交瘤技术建立了9株分泌抗IBDV VP2蛋白的单克隆抗体(Monoclonal Antibodies, mAbs)杂交瘤细胞系。间接ELISA检测9株mAbs的细胞培养上清液效价为4×10-2～1.6×10-3,腹水效价为10-2～10-5。Western-blotting结果表明,其中4株mAbs在蛋白分子量约53ku处有特异性条带,5株则未显示蛋白带。相加ELISA结果显示,单克隆抗体A12G、B12F、F9C对应相同的抗原表位,其余6株针对不同抗原表位。在夹心ELISA中,9株mAbs与新城疫病毒、传染性支气管炎病毒、减蛋综合征病毒、禽流感病毒H9N2亚型、Sf9细胞培养上清液和CEF裂解物均无交叉反应。将9株杂交瘤细胞系进行传代培养、液氮冻存与复苏实验,杂交瘤细胞仍保持稳定分泌单克隆抗体的能力。用本实验制备的mAbs建立的IBDV夹心ELISA检测方法具有良好的特异性。
Infectious bursal disease virus (IBDV), the etiological agent of infectious disease, destructs lymphatic tissue, especially the lymphocytes in the bursa of Fabricius in young chickens. It causes death and immunodepression, and results in immunity decrease and vaccination failure. This disease continues to pose significant losses in the commercial poultry industry. The epidemic situation of IBD is complex, while the vIBDV (virulent IBDV) and vvIBDV (very virulent IBDV) appeare for the past few years which can escape from common vaccinal immounization, because of the virulence and antigencity changing. IBDV VP2 protein is the main structural protein and host protective antigen, therefore, studies on VP2 protein and the detection methods for VP2 are important works at present. In this study, we have expressed the prevalent IBDV VP2 gene in Escherichia coli and baculovirus expression system respectively. Nine monoclonal antibodies (mAbs) against VP2 protein were developed. The thesis includes 3 parts:
1. Expression in Escherichia coli of VP2 gene of infectious bursal disease virus
A VP2 gene of infectious bursal disease virus (IBDV) was amplified from bursa of Fabricius of chickens with immunoprophylaxis defeat in Anhui province and cloned into the prokaryotic expression vector pGEX-4T-1. The expression plasmid pGEX-VP2 was constructed and transformed into competent Escherichia coli Rosetta (DE3), the target protein was expressed under the induction with IPTG and was positive in the detection of IBDV sandwich ELISA. SDS-PAGE analysis showed that the VP2 fusion protein was approximately 68 ku in molecular weight, and it made up 16.0% of the total bacterial proteins as inclusion bodies in E. coli. Western-blotting showed that the VP2 protein could react with IBDV antibody, indicating that the VP2 protein possessed good antigenicity.
2. Expression of VP2 gene of infectious bursal disease virus in insect cells
To develop the recombinant VP2 protein of the current infectious bursal disease virus (IBDV), the VP2 gene of IBDV isolated from chicken with immunoprophylaxis defeat in Anhui province was cloned into pFastBacHTA donor plasmid. The recombinant donor plasmid pFastBacHT-VP2 containing VP2 was constructed and transformed into competent E. coli DH10Bac cells. After screening, the recombinant expression Bacmid rBacHT-VP2 was obtained and used to transfect insect Sf9 cells with Lipofectamine reagent to produce recombinant baculovirus vBacHT-VP2. The protein band of approximate 53 ku was detected in western blotting, the Sf9 cells infected with vBac-VP2 could generate specific fluorescent light in the indirect immunofluorescence assay (IFA), and the self-assembly virus-like particles (VLPs) could be observed by electron microscopy.
3. Preparation and application of monoclonal antibodies against VP2 protein of infectious bursal disease virus
Recombinant VP2 protein of infectious bursal disease virus (IBDV) of AH1 strain expressed by Escherichia coli and recombinant baculovirus and purified IBDV antigen were used to immunize BALB/c mice, nine hybridomas secreting anti-VP2 monoclonal antibodies(mAbs) were established by hybridoma technique. The antibodies titers were 4×10-2～1.6×10-3 in culture supernatants and 10-2～10-5 in ascetic fluids by indirect-ELISA. The recombinant VP2 protein could react with four mAbs and the specific protein bands were 53 ku in western-blotting. Additivity ELISA indicated that A12G、B12F and F9C were corresponding to the same antigenic epitope, and the other six mAbs were different. The nine mAbs were specific to IBDV and didn't react with NDV, IBV, EDAV, AIV H9N2, culture supernatant of Sf9 cells and lysate of CEF in sandwich ELISA. The stability experiment indicated that the nine mAbs posed the ability of secreting antibodies after subculturing, freezing and thawing test. The sandwich ELISA established by the mAbs for detecting IBDV showed highly specificity.
1.传染性法氏囊病病毒VP2基因在大肠杆菌中的表达
将从安徽省传染性法氏囊病(IBD)免疫失败的病鸡法氏囊组织中扩增的VP2基因定向克隆入原核表达载体pGEX-4T-1中,构建了原核表达质粒pGEX-VP2,并转化大肠杆菌Rosetta (DE3).经IPTG诱导后,用IBDV双抗体夹心ELISA检测,结果呈阳性反应：SDS-PAGE分析结果显示,重组VP2融合蛋白的分子质量为68 ku,表达量约占菌体总蛋白量的16.0%,表达产物主要以包涵体形式存在于菌体内；Western-blotting结果显示,重组VP2蛋白与IBDV抗体发生反应,形成一条特异性蛋白带,表明获得的重组蛋白具有良好的抗原反应性。
2.传染性法氏囊病病毒VP2基因在昆虫细胞中的表达
为制备针对传染性法氏囊病病毒(IBDV)流行毒株的重组VP2蛋白,将从安徽省IBD免疫失败的病鸡法氏囊组织中扩增的VP2基因插入到杆状病毒表达系统供体质粒pFastBacHTA中,转化大肠杆菌DH10Bac感受态细胞,并成功获得重组杆状病毒表达质粒rBacHT-VP2,用脂质体法转染Sf9细胞,获得重组杆状病毒vBacHT-VP2.经Western-blotting分析,在蛋白分子量约53 ku处出现特异性蛋白条带；以间接免疫荧光试验(IFA)检测vBacHT-VP2感染的Sf9细胞,具有特异性免疫荧光；电镜观察,重组VP2蛋白能够自组装成病毒样颗粒(VLPs)。
3.传染性法氏囊病病毒VP2蛋白单克隆抗体的制备与应用
用大肠杆菌表达的传染性法氏囊病病毒(IBDV)重组VP2蛋白、杆状病毒表达的重组VP2蛋白以及纯化的IBDV分别免疫BALB/c小鼠,利用淋巴细胞杂交瘤技术建立了9株分泌抗IBDV VP2蛋白的单克隆抗体(Monoclonal Antibodies, mAbs)杂交瘤细胞系。间接ELISA检测9株mAbs的细胞培养上清液效价为4×10-2～1.6×10-3,腹水效价为10-2～10-5。Western-blotting结果表明,其中4株mAbs在蛋白分子量约53ku处有特异性条带,5株则未显示蛋白带。相加ELISA结果显示,单克隆抗体A12G、B12F、F9C对应相同的抗原表位,其余6株针对不同抗原表位。在夹心ELISA中,9株mAbs与新城疫病毒、传染性支气管炎病毒、减蛋综合征病毒、禽流感病毒H9N2亚型、Sf9细胞培养上清液和CEF裂解物均无交叉反应。将9株杂交瘤细胞系进行传代培养、液氮冻存与复苏实验,杂交瘤细胞仍保持稳定分泌单克隆抗体的能力。用本实验制备的mAbs建立的IBDV夹心ELISA检测方法具有良好的特异性。
Infectious bursal disease virus (IBDV), the etiological agent of infectious disease, destructs lymphatic tissue, especially the lymphocytes in the bursa of Fabricius in young chickens. It causes death and immunodepression, and results in immunity decrease and vaccination failure. This disease continues to pose significant losses in the commercial poultry industry. The epidemic situation of IBD is complex, while the vIBDV (virulent IBDV) and vvIBDV (very virulent IBDV) appeare for the past few years which can escape from common vaccinal immounization, because of the virulence and antigencity changing. IBDV VP2 protein is the main structural protein and host protective antigen, therefore, studies on VP2 protein and the detection methods for VP2 are important works at present. In this study, we have expressed the prevalent IBDV VP2 gene in Escherichia coli and baculovirus expression system respectively. Nine monoclonal antibodies (mAbs) against VP2 protein were developed. The thesis includes 3 parts:
1. Expression in Escherichia coli of VP2 gene of infectious bursal disease virus
A VP2 gene of infectious bursal disease virus (IBDV) was amplified from bursa of Fabricius of chickens with immunoprophylaxis defeat in Anhui province and cloned into the prokaryotic expression vector pGEX-4T-1. The expression plasmid pGEX-VP2 was constructed and transformed into competent Escherichia coli Rosetta (DE3), the target protein was expressed under the induction with IPTG and was positive in the detection of IBDV sandwich ELISA. SDS-PAGE analysis showed that the VP2 fusion protein was approximately 68 ku in molecular weight, and it made up 16.0% of the total bacterial proteins as inclusion bodies in E. coli. Western-blotting showed that the VP2 protein could react with IBDV antibody, indicating that the VP2 protein possessed good antigenicity.
2. Expression of VP2 gene of infectious bursal disease virus in insect cells
To develop the recombinant VP2 protein of the current infectious bursal disease virus (IBDV), the VP2 gene of IBDV isolated from chicken with immunoprophylaxis defeat in Anhui province was cloned into pFastBacHTA donor plasmid. The recombinant donor plasmid pFastBacHT-VP2 containing VP2 was constructed and transformed into competent E. coli DH10Bac cells. After screening, the recombinant expression Bacmid rBacHT-VP2 was obtained and used to transfect insect Sf9 cells with Lipofectamine reagent to produce recombinant baculovirus vBacHT-VP2. The protein band of approximate 53 ku was detected in western blotting, the Sf9 cells infected with vBac-VP2 could generate specific fluorescent light in the indirect immunofluorescence assay (IFA), and the self-assembly virus-like particles (VLPs) could be observed by electron microscopy.
3. Preparation and application of monoclonal antibodies against VP2 protein of infectious bursal disease virus
Recombinant VP2 protein of infectious bursal disease virus (IBDV) of AH1 strain expressed by Escherichia coli and recombinant baculovirus and purified IBDV antigen were used to immunize BALB/c mice, nine hybridomas secreting anti-VP2 monoclonal antibodies(mAbs) were established by hybridoma technique. The antibodies titers were 4×10-2～1.6×10-3 in culture supernatants and 10-2～10-5 in ascetic fluids by indirect-ELISA. The recombinant VP2 protein could react with four mAbs and the specific protein bands were 53 ku in western-blotting. Additivity ELISA indicated that A12G、B12F and F9C were corresponding to the same antigenic epitope, and the other six mAbs were different. The nine mAbs were specific to IBDV and didn't react with NDV, IBV, EDAV, AIV H9N2, culture supernatant of Sf9 cells and lysate of CEF in sandwich ELISA. The stability experiment indicated that the nine mAbs posed the ability of secreting antibodies after subculturing, freezing and thawing test. The sandwich ELISA established by the mAbs for detecting IBDV showed highly specificity.
引文
[1]Hermann M, Md R I, Rudiger R. Research on infectious bursal disease the past, the present and the future [J]. Veterinary Microbiology,2003,97 (1-2):153-165.
[2]Cosgrove A S. An apparently new disease of chicken-aviannephrosis [J]. Avian Dis,1962,6: 385-389.
[3]Jackwood D H, Saif Y M. Antigenic Diversity of infectious bursal disease viruses [J]. Avian Dis, 1987,31 (4):766-770.
[4]朱爱国,李劲松,陈溥言,等.鸡传染性法氏囊病毒血清亚型的研究[J].中国畜禽传染病,1992,4：1-4.
[5]李树根,黄生,林志雄,等.鸡传染性法氏囊血清Ⅰ型病毒亚型毒株的分离[J].中国畜禽传染病,1991,5：7-11.
[6]Cao Y C, Yeung W S, Law M, et al. Molecular characterization of seven Chinese isolates of infectious bursal disease virus:classical, very virulent, and variant strains [J]. Avian Dis,1998,42 (2):340-351.
[7]周宗安,王永山,邓小昭,等.传染性法氏囊病病毒的生态学与流行病学研究[J].中国兽医学报,1998,18(5)：430-433.
[8]Mundt E, Kollner B, Kretzschmar D. VP5 of infectious bursal disease virus is not essential for virus replication in cell culture [J]. J Virus,1997,71 (7):5647-5651.
[9]Morgan M M, Macreadie I G, Harley V R, et al. Sequence of the small double-stranded RNA genomic segment of infectious bursal disease virus and its deduced 90kDa product [J]. Virology, 1988,163 (1):240-242.
[10]刘爵,刘有昌,曹永长等.鸡传染性法氏囊病病毒的分子生物研究进展[J].中国兽医学报,1999,20(2)：205-208.
[11]Kibenge F S, Qian B, Cleghorn J R, et al. Infectious bursal disease virus polyprotein processing does not involve cellular proteases [J]. Arch Virol,1997,142 (12):2401-2419.
[12]Kibenge F S, Dhama V. Evidence that virion-associated VP1 of avibirnaviruses contains viral RNA sequences [J]. Arch Virol,1997,142 (6):1227-1236.
[13]Lombardo E, Maraver A, Espinosa I, et al. VP5, the non-structural polypeptide of infectious bursal disease virus, accumulates within the host plasma membrane and induces cell lysis [J]. Virology, 2000,277 (2):345-357.
[14]Lombardo E, Maraver A, Caston J R, et al. VP1, the putative RNA-dependent RNA polymerase of infectious burial disease virus, forms complexes with the capsid protein VP3, leading to efficient encapsidation into virus-like particles [J]. J Virol,1999,73 (8):6973-6983.
[15]Bayliss C D, Spies U, Shaw K, et al. A comparison of the sequences of segment A of four infectious bursal disease virus strains and identification of a variable region in VP2 [J]. J Gen Virol,1990,71 (6):1303-1312.
[16]Ikuta N, EI-Attrache J, Villegas P, et al. Molecular characterization of Brazilian infectious busral disease viruses [J]. Avian Disease,2001,45 (2):297-306.
[17]Fernandez-Arias A, Martines S, Rodriguez J F, et al. The major antigenic protein of infectious bursal disease virus, VP2, is an apoptotic inducer [J]. J Virol,1997,71 (10):8014-8018.
[18]Oppling V, Muller H, Becht H. Heterogeneity of the antigenic site responsible for the induction of neutralizing antibodies in infectious bursal disease virus [J]. Arch Virol,1991,119 (3-4):211-223.
[19]Tacken M G, Rottier P J, Gielkens A L, et al. Interactions in vivo between the proteins of infectious bursal disease virus:capsid protein VP3 interacts with the RNA-dependent RNA polymerase, VP1 [J]. J Gen Virol,2000,81(1):209-218.
[20]Martinez-Torrecuadrada J L, Caston J R, Castro M, et al. Different architectures in the assembly of infectious bursal disease virus capsid proteins expressed in insect cells [J]. Virology,2000,278 (2):322-331.
[21]Lejal N, Da Costa B, Huet J C, et al. Role of Ser-652 and Lys-692 in the protease activity of infectious bursal disease virus VP4 and identification of its substrate cleavage sites [J]. J Gen Virol, 2000,81 (9):983-992.
[22]Granzow H, Bigrhan C, Mettenleiter T C, et al. A second form of infectious bursal disease virus-associated tubule contains VP4 [J]. J Virol,1997,71 (11):8879-8885.
[23]Yao K, Vakharia V N. Induction of apoptosis in vitro by the 17-kDa nonstructural protein of infectious bursal disease virus:possible role in viral pathogenesis [J]. Virology,2001,285 (1): 50-58.
[24]白文彬,于康震.动物传染病诊断学[M].北京：中国农业出版社,2002：238-245.
[25]马兴树,辛盛鹏,阎志民,等.SPA直接平板协同凝集试验诊断鸡传染性法氏囊病[J].中国兽医学报,2002,22(2)：116-117.
[26]樊晓京,朱益群,黄建生,等.用反向间接血凝抑制试验检测传染性法氏囊病血清抗体[J].中国畜禽传染病,1997,3：31-33.
[27]王桂枝,毕丁仁,陶秀萍,等.应用双抗体夹心法ELISA检测IBDV的研究[J].中国兽医杂志,1997,20(7)：3-5.
[28]马兴树,叶树华.应用火箭免疫电泳技术诊断鸡传染性法氏囊病的研究[J].中国畜禽传染病, 1995,81(2)：17-18.
[29]Kusk M, Kabell S, Jorgensen P H, et al. Differentiation of five strains of infectious bursal disease virus:development of a strain-specific multiplex PCR [J]. Vet Microbiol,2005,109 (3-4):159-167.
[30]Tiwari A K, Kataria R S, Indervesh, et al. Differentiation of infectious bursal disease viruses by restriction enzyme analysis of RT-PCR amplified VP1 gene sequence [J].Comp Immunol Microbio Infect Dis,2003,26 (1):47-53.
[31]陈士友,陈溥言,蔡宝祥,等.应用DIG标记探针杂交检测IBDV[J].中国病毒学,1994,9(4)：351-355.
[32]王忠灿,王永山,唐雨德,等.传染性法氏囊病病毒检测系统的建立[J].中国兽医学报,2008,28(9)：1015-1019.
[33]Azad A A, McKern N M, Macreadie I G, et al. Physicochemical and immunological characterization of recombinant host-protective antigen(VP2) of infectious bursal disease virus[J]. Vaccine,1991,9 (10):715-722.
[34]Azad A A, Fahey K J, Barrett S A, et al. Expression in Escherichia coli of cDNA fragments encoding the gene for the host-protective antigen of infectious bursal disease virus [J]. Virology, 1986,149 (1):190-198.
[35]姜平,陈溥言,蔡宝祥.IBDV南京野毒株VP2结构蛋白基因克隆与表达[J].南京农业大学学报,1996,19(4)：62-65.
[36]曹冰玉,王永山,范红结,等.引起免疫失败的传染性腔上囊病病毒VP2基因在大肠杆菌中的表达[J].中国兽医科学,2010,40(2)：130-134.
[37]Macreadie I G, Vaughan P R, Chapman A J, et al. Passive protection against infectious busral disease virus by viral VP2 experssed in yeast [J]. Vaccine,1990,8 (6):549-552.
[38]邓继先.昆虫杆状病毒载体细胞培养和重组蛋白生产[J].生物工程进展,1993,13：53-56.
[39]宋坤华,金勇丰,黄耀伟,等.重组家蚕病毒表达传染性法氏囊病病毒VP2蛋白[J].生物化学与生物物理学报,2000,32(3)：281-284.
[40]于涟,宋坤华,张耀洲,等.家蚕表达传染性法氏囊病病毒VP2蛋白的免疫原性初步研究[J].浙江大学学报(农业与生命科学版),2000,26(1)：9-16.
[41]Bayliss C D, Peters R W, Cook J K, et al. A recombinant fowlpox virus that expresses the VP2 antigen of infectious bursal disease virus induces protection against mortality caused by the virus [J]. Arch Virol,1991,120 (3-4):193-205.
[42]Sheppard M, Werner W, Tsatas D, et al. Fowl adenovirus recombinant expressing VP2 of infectious bursal disease virus induces protective immunity against bursal disease [J]. Arch Virol,1998,142 (5):915-930.
[43]Darteil R, Bublot M, Laplace E, et al. Herpesvirus of turkey recombinant viruses expressing infectious bursal disease virus (IBDV) VP2 immunogen induce protection against an IBDV virulent challenge in chickens [J]. Virology,1995,211 (2):481-490.
[44]Tsukamoto K, Kojima C, Komori Y, et al. Protection of chickens against very infectious bursal disease virus(IBDV) and Marek's disease(MDV) with a recombinant MDV expressing IBDV virus VP2 [J]. Virology,1999,257 (2):352-362.
[45]Fordor N, Dube S K, Fodor I, et al. Induction of protective immunity in chickens immunized with plasmid DNA encoding infectious bursal disease virus antigens [J]. Acta Vet Hung,1999,47 (4):481-492.
[46]Mahmood M S, Hussain I, Siddique M, et al. DNA vaccination with VP2 gene of very virulent infectious bursal disease virus(wIBDV) delivered by transgenic E. coli DH5a given orally confers protective immune responses in chickens [J]. Vaccine,2007,25 (44):7629-7635.
[47]Meyer D, Torres JV. Hypervariable epitope construct:a synthetic immunogen that overcomes MHC restriction of antigen presentation [J]. Mol Immunol,1999,36 (10):631-637.
[48]Ulrich R, Nassal M, Meisel H, et al. Core particles of hepatitis B virus as carrier for foreign epitopes [J]. Adv Virus Res,1998,50:141-182.
[49]Ohara N, Matsuoka M, Nomaguchi H, et al. Protective responses against experimental Mycobacterium leprae infection in mice induced by recombinant Bacillus Calmette-Guerin over-producing three putative protective antigen candidates [J]. Vaccine,2001,19 (15-16):1906-1910.
[50]Ravn P, Demissie A, Eguale T, et al. Human T cell responses to the ESAT-6 antigen from Mycobacterium tuberculosis [J]. J Infect Dis,1999,179 (3):637-645.
[51]刘小娟,王永山,欧阳伟,等.传染性法氏囊病病毒抗原表位与乙型肝炎病毒核心抗原嵌合基因的构建及其表达产物分析[J].中国预防兽医学报,2010,32(11)：879-883.
[52]王忠灿,王永山,刘小娟,等.IBDV抗原表位与HBcAg嵌合基因的分子设计.中国,专利申请号：200910232402.3
[53]王廷华,李官成,Zhou X F.抗体理论与技术[M].第2版.北京：科学出版社,2005：91-92.
[54]刘秀梵.单克隆抗体在农业上的应用[M].合肥：安徽科技出版社,1994：47-59.
[55]Kufer P, Lutterbuse R, Baeuerle P A. The promise of bispecific antibodies [J]. Discov Med,2004,4 (23):325-332.
[56]Siddiqui M Z. Monoclonal antibodies as diagnostics; an appraisal [J]. Indian J Pharm Sci,2010,72 (1):12-17.
[57]Weiner, Louis M. Fully human therapeutic monoclonal antibodies [J]. J Immunother,2006,29 (1):1-9.
[58]Yokota T, Milenic D E, Whitlow M, et al. Rapid tumor penetration of a single-chain Fv and comparison with other immunoglobulin forms [J]. Cancer Res,1992,52 (12):3402-3408.
[59]Benhar I, Reiter Y, Pai L H, et al. Administration of disulfide-stabilized Fv-immunoloxins B1(dsFv)-PE38 by continuous infusion increases their efficacy in curing large tumor xenografts in nude mice [J]. Int J Cancer,1995,62 (3):351-355.
[60]朱红.用抗传染性法氏囊病毒单克隆抗体建立夹心阻断ELISA检测鸡血清抗体及初步应用[J].动物科学与动物医学,2000,17(5)：51-52.
[61]Zhang G P, Li Q M, Yang Y Y, et al. Development of a one-step strip test for the diagnosis of chicken infectious bursal disease [J]. Avian Dis,2005,49 (2):177-181.
[62]Yamaguchi T, Lwata K, Kobayashi M, et al. Epitope mapping of capsid proteins VP2 of infectious bursal disease virus [J]. Arch Virol,1996,141 (8):1493-1507.
[63]Cui X, Nagesha H S, Holmes I H. Mapping of conformational epitopes on capsid protein VP2 of infectious bursal disease virus by fd-tet phage display [J]. J Virol Methods,2003,114 (l):190-112.
[64]Wang X N, Zhang G P, Zhou J Y, et al. Identification of neutralizing epitopes on the VP2 protein of infectious bursal disease virus by phage-displayed heptapeptide library screening and synthetic peptide mapping [J]. Viral Immunol,2005,18 (3):549-557.
[65]王军,李银,范红结,等.传染性法氏囊病病毒抗原表位分析-单克隆抗体的制备与鉴定[J].中国预防兽医学报,2005,27(3)：171-174.
[66]金山,倪龙根,王永坤.抗鸡传染性法氏囊病病毒型特异性单克隆抗体的临床治疗试验[J].中国畜禽传染病,1997,19(6)：57-58.
[1]Muller H, Islam M R, Raue R. Research on infectious bursal disease—the past, the present and the future [J]. Vet Microbiol,2003,97 (1-2):153-165.
[2]Wang Y S, Wang Z C, Tang Y D, et al. Comparison of four infectious bursal disease viruses isolated from different bird species [J]. Arch Virol,2007,152 (10):1787-1797.
[3]Cao Y C, Yeung W S, Law M, et al. Molecular characterization of seven Chinese isolates of infectious bursal disease virus:classical, very virulent and variant strains [J]. Avian Dis,1998,42 (2):340-351.
[4]Ymaguchi T, Ogawa M, Miyoshi M, et al. Sequence and phylogenetic analyses of highly virulent infectious bursal disease virus [J]. Arch Virol,1997,142 (7):1441-1458.
[5]Maradassi H, Khabouchi N, Ghram A, et al. A very virulent genotype of infectious bursal disease virus predominantly associated with recurrent infectious bursal disease outbreaks in Tunisian vaccinated flocks [J]. Avian Dis,2004,48 (4):829-840.
[6]Yamaguchi T, Kasanga C J, Terasaki K, et al. Nucleotide sequence analysis of VP2 hypervariable domain of infectious bursal disease virus detected in Japan from 1993 to 2004 [J]. J Vet Med Sci, 2007,69 (7):733-738.
[7]Dola R, Majo N, Ordonez G, et al. Viral genotyping of infectious bursal disease viruses isolated from the 2002 acute outbreak in Spain and comparison with previous isolates [J]. Avian Dis,2005, 49 (3):332-339.
[8]王永山,欧阳伟,潘群兴,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因的分子特征[J].中国兽医科学,2008,38(12)：919-925.
[9]Lombardo E, Maraver A, Espinosa I, et al. VP5, the nonstructural polypeptide of infectious bursal disease virus, accumulates within the host plasma membrane and induces cell lysis [J]. Virology, 2000,277 (2):345-357.
[10]祁小乐,王笑梅,高玉龙,等.鸡传染性法氏囊病病毒VP2蛋白研究进展[J].中国预防兽医学报,2008,30(8)：656-660.
[11]Azad A A, Fahey K J, Barrett S A, et al. Expression in Escherichia coli of cDNA fragments encoding the gene for the host-protective antigen of infectious bursal disease virus [J]. Virology, 1986,149 (2):190-198.
[12]Macreadie I G, Vaugham P R, Chapman A J, et al. Passive protection against infectious bursal disease virus by viral VP2 expressed in yeast [J]. Vaccine,1990,8 (6):549-552.
[13]Shaw I, Davison T F. Protection from IBDV-induced bursal damage by a recombinant fowlpox vaccine, fpIBD1, is dependent on the titer of challenge virus and chicken genotype [J]. Vaccine, 2000,18(28):3230-3241.
[14]Snyder D B, Vakharia V N, Mengel-Whereat S A, et al. Active cross-protection induced by a recombinant baculovirus expressing chimeric infectious bursal disease virus structural proteins[J]. Avian Dis,1994,38 (4):701-707.
[15]Fodor I, Horvath E, Fodor N, et al. Induction of protective immunity in chickens immunized with plasmid DNA encoding infectious bursal disease virus antigens [J]. Acta Vet Hung,1999,47 (4):481-492.
[16]Wang Y S, Fan H J, Li Y, et al. Development of a multi-mimotope peptide as a vaccine immunogen for infectious bursal disease virus [J]. Vaccine,2007,25 (22):4447-4455.
[17]Ahn S J, Seo J S, Kim M S, et al. Cloning, site-directed mutagenesis and expression of cathepsin L-like cysteine protease from Uronema marinum (Ciliophora:Scuticociliatida) [J]. Mol Biochem Parasit,2007,156 (2):191-198.
[18]Fu W, Lin J, Cen P.5-Aminolevulinate production with recombinant Escherichia coli using a rare codon optimizer host strain [J]. Appl Microbiol Biot,2007,75 (4):777-782.
[19]Rabhi-Essafi I, Sadok A, Khalaf N, et al. A strategy for high-level expression of soluble and functional human interferon alpha as a GST-fusion protein in E. coli [J]. Protein Eng Des Sel, 2007,20(5):201-209.
[20]吴静,雷楗勇,张莲芬,等.改善稀有密码子和氨基酸残基限制提高重组人ADAM15去整合素结构域蛋白表达水平[J].微生物学报,2008,48(8)：1067-1074.
[21]Armando F A, Siomara M, Jose F R. The major antigenic protein of infectious bursal disease virus, VP2, is an apoptotic inducer [J]. J Virol,1997,71 (10):8014-8018.
[1]Maradassi H, Khabouchi N, Ghram A, et al. A very virulent genotype of infectious bursal disease virus predominantly associated with recurrent infectious bursal disease outbreaks in Tunisian vaccinated flocks [J]. Avian Dis,2004,48 (4):829-840.
[2]Wang Y S, Wang Z C, Tang Y D, et al. Comparison of four infectious bursal disease viruses isolated from different bird species [J]. Archives of Virology,2007,152 (10):1787-1797.
[3]Muller H, Islam M R, Raue R, et al. Research on infectious bursal disease—the past, the present and the future [J]. Vet Microbiol,2003,97 (1-2):153-165.
[4]Yamaguchi T, Iwata K, Kobayashi M, et al. Epitope mapping of capsid proteins VP2 and VP3 of infectious bursal disease virus [J]. Arch Virol,1996,141 (8):1493-1507.
[5]王永山,欧阳伟,潘群兴,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因的分子特征[J].中国兽医科学,2008,38(12)：919-925.
[6]Fahey K J, Erny K, Crooks J, et al. A conformational immunogen on VP2 of infectious bursal disease virus that induces virus-neutralizing antibodies that passively protect chickens [J]. J Gen Virol,1989,70(6):1473-1481.
[7]Heine H G, Haritou M, Failla P, et al. Sequence analysis and expression of the host-protective immunogen VP2 of a variant strain of infectious bursal disease virus which can circumvent vaccination with standard type I strains [J]. J Gen Virol,1991,72(8):1835-1843.
[8]曹冰玉,王永山,范红结,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因在大肠杆菌中的表达[J].中国兽医科学,2010,40(2)：130-134.
[9]Ulrich R, Nassal M, Meisel H, et al. Core particles of hepatitis B virus as carrier for foreign epitopes [J].Adv Virus Res,1998,50:141-182.
[10]Prel A, Le Gall-Recule G, Jestin V. Achievement of avian influenza virus-like particles that could be used as a subunit vaccine against low-pathogenic avian influenza strains in ducks [J]. Avian Pathol,2008,37 (5):513-520.
[11]Cui X, Nagesha H S, Holmes I H. Mapping of conformational epitopes on capsid protein VP2 of infectious bursal disease virus by fd-tet phage display [J]. J Virol Methods,2003,114 (1):109-112.
[1]卡尔尼克B w主编.禽病学[M].第10版.高福,苏敬良,主译.北京：中国农业出版社,1999：914-937.
[2]Muller H, Islam M R, Raue R. Research on infectious bursal disease—the past, the present and the future [J]. Vet Microbiol,2003,97 (1-2):153-165.
[3]Kibenge F S, Jackwood D J, Mercado C C. Nucleotide sequence analysis of genome segment A of infectious bursal disease virus [J]. J Gen Virol,1990,71 (3):569-577.
[4]Yamaguchi T, Lwata K, Kobayashi M, et al. Epitope mapping of capsid proteins VP2 and VP3 of infectious bursal disease virus [J]. Arch Virol,1996,141 (8):1493-1507.
[5]Heine H G, Hariton M, Failla P, et al. Sequence analysis and expression of the host-protective immunogen VP2 of a variant strain of infectious bursal disease virus which can circumvent vaccination with standard type I strains [J]. J Gen Virol,1991,72 (8):1835-1843.
[6]Cui X, Nagesha H S, Holmes I H. Mapping of conformational epitopes on capsid protein VP2 of infectious bursal disease virus by fd-tet phage display [J]. J Virol Methods,2003,114 (1):109-112.
[7]Yao K, Vakharia V N. Induction of apoptosis in vitro by the 17-kDa nonstructural protein of infectious bursal disease virus:possible role in viral pathogenesis [J]. Virology,2001,285 (1):50-58.
[8]王永山,范红结,李银,等.传染性法氏囊病病毒五个抗原表位短肽的鉴定与序列分析[J].中国病毒学,2005,20(5)：503-506.
[9]祁小乐,王笑梅,高玉龙,等.鸡传染性法氏囊病病毒VP2蛋白研究进展[J].中国预防兽医学报,2008,30(8)：656-660.
[10]王永山,欧阳伟,潘群兴,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因的分子特征[J].中国兽医科学,2008,38(12)：919-925.
[11]曹冰玉,王永山,范红结,等.引起免疫失败的传染性腔上囊病病毒VP2基因在大肠杆菌中的表达[J].中国兽医科学,2010,40(2)：130-134.
[12]王军,李银,范红结,等.传染性法氏囊病病毒抗原表位分析一单克隆抗体的制备与鉴定[J].中国预防兽医学报,2005,27(3)：171-174.
[13]王永山,郝崇,侯世宽,等.抗鸡IgG单克隆抗体的研究[J].兽医大学学报,1989,9(2)：109-114.
[14]余抒,罗萍,陈洪章,等.抗肠出血性大肠杆菌O157:H7 EspA单克隆抗体的制备及其特异性鉴定[J].细胞与分子免疫学杂志,2007,23(7)：657-659.
[15]刘玉斌,苟仕金,主编.动物免疫学实验技术[M].吉林：吉林科学技术出版社,1989：150-163.
[16]Wang X N, Zhang G P, Zhou J Y, et al. Identification of neutralizing epitopes on the VP2 protein of infectious bursal disease virus by phage-Displayed heptapeptide library screening and synthetic peptide [J]. Viral Immunology,2005,18 (3):549-557.
[17]王忠灿,王永山,唐雨德,等.传染性法氏囊病病毒检测系统的建立[J].中国兽医学报,2008,28(9)：1015-1019.
[2]Cosgrove A S. An apparently new disease of chicken-aviannephrosis [J]. Avian Dis,1962,6: 385-389.
[3]Jackwood D H, Saif Y M. Antigenic Diversity of infectious bursal disease viruses [J]. Avian Dis, 1987,31 (4):766-770.
[4]朱爱国,李劲松,陈溥言,等.鸡传染性法氏囊病毒血清亚型的研究[J].中国畜禽传染病,1992,4：1-4.
[5]李树根,黄生,林志雄,等.鸡传染性法氏囊血清Ⅰ型病毒亚型毒株的分离[J].中国畜禽传染病,1991,5：7-11.
[6]Cao Y C, Yeung W S, Law M, et al. Molecular characterization of seven Chinese isolates of infectious bursal disease virus:classical, very virulent, and variant strains [J]. Avian Dis,1998,42 (2):340-351.
[7]周宗安,王永山,邓小昭,等.传染性法氏囊病病毒的生态学与流行病学研究[J].中国兽医学报,1998,18(5)：430-433.
[8]Mundt E, Kollner B, Kretzschmar D. VP5 of infectious bursal disease virus is not essential for virus replication in cell culture [J]. J Virus,1997,71 (7):5647-5651.
[9]Morgan M M, Macreadie I G, Harley V R, et al. Sequence of the small double-stranded RNA genomic segment of infectious bursal disease virus and its deduced 90kDa product [J]. Virology, 1988,163 (1):240-242.
[10]刘爵,刘有昌,曹永长等.鸡传染性法氏囊病病毒的分子生物研究进展[J].中国兽医学报,1999,20(2)：205-208.
[11]Kibenge F S, Qian B, Cleghorn J R, et al. Infectious bursal disease virus polyprotein processing does not involve cellular proteases [J]. Arch Virol,1997,142 (12):2401-2419.
[12]Kibenge F S, Dhama V. Evidence that virion-associated VP1 of avibirnaviruses contains viral RNA sequences [J]. Arch Virol,1997,142 (6):1227-1236.
[13]Lombardo E, Maraver A, Espinosa I, et al. VP5, the non-structural polypeptide of infectious bursal disease virus, accumulates within the host plasma membrane and induces cell lysis [J]. Virology, 2000,277 (2):345-357.
[14]Lombardo E, Maraver A, Caston J R, et al. VP1, the putative RNA-dependent RNA polymerase of infectious burial disease virus, forms complexes with the capsid protein VP3, leading to efficient encapsidation into virus-like particles [J]. J Virol,1999,73 (8):6973-6983.
[15]Bayliss C D, Spies U, Shaw K, et al. A comparison of the sequences of segment A of four infectious bursal disease virus strains and identification of a variable region in VP2 [J]. J Gen Virol,1990,71 (6):1303-1312.
[16]Ikuta N, EI-Attrache J, Villegas P, et al. Molecular characterization of Brazilian infectious busral disease viruses [J]. Avian Disease,2001,45 (2):297-306.
[17]Fernandez-Arias A, Martines S, Rodriguez J F, et al. The major antigenic protein of infectious bursal disease virus, VP2, is an apoptotic inducer [J]. J Virol,1997,71 (10):8014-8018.
[18]Oppling V, Muller H, Becht H. Heterogeneity of the antigenic site responsible for the induction of neutralizing antibodies in infectious bursal disease virus [J]. Arch Virol,1991,119 (3-4):211-223.
[19]Tacken M G, Rottier P J, Gielkens A L, et al. Interactions in vivo between the proteins of infectious bursal disease virus:capsid protein VP3 interacts with the RNA-dependent RNA polymerase, VP1 [J]. J Gen Virol,2000,81(1):209-218.
[20]Martinez-Torrecuadrada J L, Caston J R, Castro M, et al. Different architectures in the assembly of infectious bursal disease virus capsid proteins expressed in insect cells [J]. Virology,2000,278 (2):322-331.
[21]Lejal N, Da Costa B, Huet J C, et al. Role of Ser-652 and Lys-692 in the protease activity of infectious bursal disease virus VP4 and identification of its substrate cleavage sites [J]. J Gen Virol, 2000,81 (9):983-992.
[22]Granzow H, Bigrhan C, Mettenleiter T C, et al. A second form of infectious bursal disease virus-associated tubule contains VP4 [J]. J Virol,1997,71 (11):8879-8885.
[23]Yao K, Vakharia V N. Induction of apoptosis in vitro by the 17-kDa nonstructural protein of infectious bursal disease virus:possible role in viral pathogenesis [J]. Virology,2001,285 (1): 50-58.
[24]白文彬,于康震.动物传染病诊断学[M].北京：中国农业出版社,2002：238-245.
[25]马兴树,辛盛鹏,阎志民,等.SPA直接平板协同凝集试验诊断鸡传染性法氏囊病[J].中国兽医学报,2002,22(2)：116-117.
[26]樊晓京,朱益群,黄建生,等.用反向间接血凝抑制试验检测传染性法氏囊病血清抗体[J].中国畜禽传染病,1997,3：31-33.
[27]王桂枝,毕丁仁,陶秀萍,等.应用双抗体夹心法ELISA检测IBDV的研究[J].中国兽医杂志,1997,20(7)：3-5.
[28]马兴树,叶树华.应用火箭免疫电泳技术诊断鸡传染性法氏囊病的研究[J].中国畜禽传染病, 1995,81(2)：17-18.
[29]Kusk M, Kabell S, Jorgensen P H, et al. Differentiation of five strains of infectious bursal disease virus:development of a strain-specific multiplex PCR [J]. Vet Microbiol,2005,109 (3-4):159-167.
[30]Tiwari A K, Kataria R S, Indervesh, et al. Differentiation of infectious bursal disease viruses by restriction enzyme analysis of RT-PCR amplified VP1 gene sequence [J].Comp Immunol Microbio Infect Dis,2003,26 (1):47-53.
[31]陈士友,陈溥言,蔡宝祥,等.应用DIG标记探针杂交检测IBDV[J].中国病毒学,1994,9(4)：351-355.
[32]王忠灿,王永山,唐雨德,等.传染性法氏囊病病毒检测系统的建立[J].中国兽医学报,2008,28(9)：1015-1019.
[33]Azad A A, McKern N M, Macreadie I G, et al. Physicochemical and immunological characterization of recombinant host-protective antigen(VP2) of infectious bursal disease virus[J]. Vaccine,1991,9 (10):715-722.
[34]Azad A A, Fahey K J, Barrett S A, et al. Expression in Escherichia coli of cDNA fragments encoding the gene for the host-protective antigen of infectious bursal disease virus [J]. Virology, 1986,149 (1):190-198.
[35]姜平,陈溥言,蔡宝祥.IBDV南京野毒株VP2结构蛋白基因克隆与表达[J].南京农业大学学报,1996,19(4)：62-65.
[36]曹冰玉,王永山,范红结,等.引起免疫失败的传染性腔上囊病病毒VP2基因在大肠杆菌中的表达[J].中国兽医科学,2010,40(2)：130-134.
[37]Macreadie I G, Vaughan P R, Chapman A J, et al. Passive protection against infectious busral disease virus by viral VP2 experssed in yeast [J]. Vaccine,1990,8 (6):549-552.
[38]邓继先.昆虫杆状病毒载体细胞培养和重组蛋白生产[J].生物工程进展,1993,13：53-56.
[39]宋坤华,金勇丰,黄耀伟,等.重组家蚕病毒表达传染性法氏囊病病毒VP2蛋白[J].生物化学与生物物理学报,2000,32(3)：281-284.
[40]于涟,宋坤华,张耀洲,等.家蚕表达传染性法氏囊病病毒VP2蛋白的免疫原性初步研究[J].浙江大学学报(农业与生命科学版),2000,26(1)：9-16.
[41]Bayliss C D, Peters R W, Cook J K, et al. A recombinant fowlpox virus that expresses the VP2 antigen of infectious bursal disease virus induces protection against mortality caused by the virus [J]. Arch Virol,1991,120 (3-4):193-205.
[42]Sheppard M, Werner W, Tsatas D, et al. Fowl adenovirus recombinant expressing VP2 of infectious bursal disease virus induces protective immunity against bursal disease [J]. Arch Virol,1998,142 (5):915-930.
[43]Darteil R, Bublot M, Laplace E, et al. Herpesvirus of turkey recombinant viruses expressing infectious bursal disease virus (IBDV) VP2 immunogen induce protection against an IBDV virulent challenge in chickens [J]. Virology,1995,211 (2):481-490.
[44]Tsukamoto K, Kojima C, Komori Y, et al. Protection of chickens against very infectious bursal disease virus(IBDV) and Marek's disease(MDV) with a recombinant MDV expressing IBDV virus VP2 [J]. Virology,1999,257 (2):352-362.
[45]Fordor N, Dube S K, Fodor I, et al. Induction of protective immunity in chickens immunized with plasmid DNA encoding infectious bursal disease virus antigens [J]. Acta Vet Hung,1999,47 (4):481-492.
[46]Mahmood M S, Hussain I, Siddique M, et al. DNA vaccination with VP2 gene of very virulent infectious bursal disease virus(wIBDV) delivered by transgenic E. coli DH5a given orally confers protective immune responses in chickens [J]. Vaccine,2007,25 (44):7629-7635.
[47]Meyer D, Torres JV. Hypervariable epitope construct:a synthetic immunogen that overcomes MHC restriction of antigen presentation [J]. Mol Immunol,1999,36 (10):631-637.
[48]Ulrich R, Nassal M, Meisel H, et al. Core particles of hepatitis B virus as carrier for foreign epitopes [J]. Adv Virus Res,1998,50:141-182.
[49]Ohara N, Matsuoka M, Nomaguchi H, et al. Protective responses against experimental Mycobacterium leprae infection in mice induced by recombinant Bacillus Calmette-Guerin over-producing three putative protective antigen candidates [J]. Vaccine,2001,19 (15-16):1906-1910.
[50]Ravn P, Demissie A, Eguale T, et al. Human T cell responses to the ESAT-6 antigen from Mycobacterium tuberculosis [J]. J Infect Dis,1999,179 (3):637-645.
[51]刘小娟,王永山,欧阳伟,等.传染性法氏囊病病毒抗原表位与乙型肝炎病毒核心抗原嵌合基因的构建及其表达产物分析[J].中国预防兽医学报,2010,32(11)：879-883.
[52]王忠灿,王永山,刘小娟,等.IBDV抗原表位与HBcAg嵌合基因的分子设计.中国,专利申请号：200910232402.3
[53]王廷华,李官成,Zhou X F.抗体理论与技术[M].第2版.北京：科学出版社,2005：91-92.
[54]刘秀梵.单克隆抗体在农业上的应用[M].合肥：安徽科技出版社,1994：47-59.
[55]Kufer P, Lutterbuse R, Baeuerle P A. The promise of bispecific antibodies [J]. Discov Med,2004,4 (23):325-332.
[56]Siddiqui M Z. Monoclonal antibodies as diagnostics; an appraisal [J]. Indian J Pharm Sci,2010,72 (1):12-17.
[57]Weiner, Louis M. Fully human therapeutic monoclonal antibodies [J]. J Immunother,2006,29 (1):1-9.
[58]Yokota T, Milenic D E, Whitlow M, et al. Rapid tumor penetration of a single-chain Fv and comparison with other immunoglobulin forms [J]. Cancer Res,1992,52 (12):3402-3408.
[59]Benhar I, Reiter Y, Pai L H, et al. Administration of disulfide-stabilized Fv-immunoloxins B1(dsFv)-PE38 by continuous infusion increases their efficacy in curing large tumor xenografts in nude mice [J]. Int J Cancer,1995,62 (3):351-355.
[60]朱红.用抗传染性法氏囊病毒单克隆抗体建立夹心阻断ELISA检测鸡血清抗体及初步应用[J].动物科学与动物医学,2000,17(5)：51-52.
[61]Zhang G P, Li Q M, Yang Y Y, et al. Development of a one-step strip test for the diagnosis of chicken infectious bursal disease [J]. Avian Dis,2005,49 (2):177-181.
[62]Yamaguchi T, Lwata K, Kobayashi M, et al. Epitope mapping of capsid proteins VP2 of infectious bursal disease virus [J]. Arch Virol,1996,141 (8):1493-1507.
[63]Cui X, Nagesha H S, Holmes I H. Mapping of conformational epitopes on capsid protein VP2 of infectious bursal disease virus by fd-tet phage display [J]. J Virol Methods,2003,114 (l):190-112.
[64]Wang X N, Zhang G P, Zhou J Y, et al. Identification of neutralizing epitopes on the VP2 protein of infectious bursal disease virus by phage-displayed heptapeptide library screening and synthetic peptide mapping [J]. Viral Immunol,2005,18 (3):549-557.
[65]王军,李银,范红结,等.传染性法氏囊病病毒抗原表位分析-单克隆抗体的制备与鉴定[J].中国预防兽医学报,2005,27(3)：171-174.
[66]金山,倪龙根,王永坤.抗鸡传染性法氏囊病病毒型特异性单克隆抗体的临床治疗试验[J].中国畜禽传染病,1997,19(6)：57-58.
[1]Muller H, Islam M R, Raue R. Research on infectious bursal disease—the past, the present and the future [J]. Vet Microbiol,2003,97 (1-2):153-165.
[2]Wang Y S, Wang Z C, Tang Y D, et al. Comparison of four infectious bursal disease viruses isolated from different bird species [J]. Arch Virol,2007,152 (10):1787-1797.
[3]Cao Y C, Yeung W S, Law M, et al. Molecular characterization of seven Chinese isolates of infectious bursal disease virus:classical, very virulent and variant strains [J]. Avian Dis,1998,42 (2):340-351.
[4]Ymaguchi T, Ogawa M, Miyoshi M, et al. Sequence and phylogenetic analyses of highly virulent infectious bursal disease virus [J]. Arch Virol,1997,142 (7):1441-1458.
[5]Maradassi H, Khabouchi N, Ghram A, et al. A very virulent genotype of infectious bursal disease virus predominantly associated with recurrent infectious bursal disease outbreaks in Tunisian vaccinated flocks [J]. Avian Dis,2004,48 (4):829-840.
[6]Yamaguchi T, Kasanga C J, Terasaki K, et al. Nucleotide sequence analysis of VP2 hypervariable domain of infectious bursal disease virus detected in Japan from 1993 to 2004 [J]. J Vet Med Sci, 2007,69 (7):733-738.
[7]Dola R, Majo N, Ordonez G, et al. Viral genotyping of infectious bursal disease viruses isolated from the 2002 acute outbreak in Spain and comparison with previous isolates [J]. Avian Dis,2005, 49 (3):332-339.
[8]王永山,欧阳伟,潘群兴,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因的分子特征[J].中国兽医科学,2008,38(12)：919-925.
[9]Lombardo E, Maraver A, Espinosa I, et al. VP5, the nonstructural polypeptide of infectious bursal disease virus, accumulates within the host plasma membrane and induces cell lysis [J]. Virology, 2000,277 (2):345-357.
[10]祁小乐,王笑梅,高玉龙,等.鸡传染性法氏囊病病毒VP2蛋白研究进展[J].中国预防兽医学报,2008,30(8)：656-660.
[11]Azad A A, Fahey K J, Barrett S A, et al. Expression in Escherichia coli of cDNA fragments encoding the gene for the host-protective antigen of infectious bursal disease virus [J]. Virology, 1986,149 (2):190-198.
[12]Macreadie I G, Vaugham P R, Chapman A J, et al. Passive protection against infectious bursal disease virus by viral VP2 expressed in yeast [J]. Vaccine,1990,8 (6):549-552.
[13]Shaw I, Davison T F. Protection from IBDV-induced bursal damage by a recombinant fowlpox vaccine, fpIBD1, is dependent on the titer of challenge virus and chicken genotype [J]. Vaccine, 2000,18(28):3230-3241.
[14]Snyder D B, Vakharia V N, Mengel-Whereat S A, et al. Active cross-protection induced by a recombinant baculovirus expressing chimeric infectious bursal disease virus structural proteins[J]. Avian Dis,1994,38 (4):701-707.
[15]Fodor I, Horvath E, Fodor N, et al. Induction of protective immunity in chickens immunized with plasmid DNA encoding infectious bursal disease virus antigens [J]. Acta Vet Hung,1999,47 (4):481-492.
[16]Wang Y S, Fan H J, Li Y, et al. Development of a multi-mimotope peptide as a vaccine immunogen for infectious bursal disease virus [J]. Vaccine,2007,25 (22):4447-4455.
[17]Ahn S J, Seo J S, Kim M S, et al. Cloning, site-directed mutagenesis and expression of cathepsin L-like cysteine protease from Uronema marinum (Ciliophora:Scuticociliatida) [J]. Mol Biochem Parasit,2007,156 (2):191-198.
[18]Fu W, Lin J, Cen P.5-Aminolevulinate production with recombinant Escherichia coli using a rare codon optimizer host strain [J]. Appl Microbiol Biot,2007,75 (4):777-782.
[19]Rabhi-Essafi I, Sadok A, Khalaf N, et al. A strategy for high-level expression of soluble and functional human interferon alpha as a GST-fusion protein in E. coli [J]. Protein Eng Des Sel, 2007,20(5):201-209.
[20]吴静,雷楗勇,张莲芬,等.改善稀有密码子和氨基酸残基限制提高重组人ADAM15去整合素结构域蛋白表达水平[J].微生物学报,2008,48(8)：1067-1074.
[21]Armando F A, Siomara M, Jose F R. The major antigenic protein of infectious bursal disease virus, VP2, is an apoptotic inducer [J]. J Virol,1997,71 (10):8014-8018.
[1]Maradassi H, Khabouchi N, Ghram A, et al. A very virulent genotype of infectious bursal disease virus predominantly associated with recurrent infectious bursal disease outbreaks in Tunisian vaccinated flocks [J]. Avian Dis,2004,48 (4):829-840.
[2]Wang Y S, Wang Z C, Tang Y D, et al. Comparison of four infectious bursal disease viruses isolated from different bird species [J]. Archives of Virology,2007,152 (10):1787-1797.
[3]Muller H, Islam M R, Raue R, et al. Research on infectious bursal disease—the past, the present and the future [J]. Vet Microbiol,2003,97 (1-2):153-165.
[4]Yamaguchi T, Iwata K, Kobayashi M, et al. Epitope mapping of capsid proteins VP2 and VP3 of infectious bursal disease virus [J]. Arch Virol,1996,141 (8):1493-1507.
[5]王永山,欧阳伟,潘群兴,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因的分子特征[J].中国兽医科学,2008,38(12)：919-925.
[6]Fahey K J, Erny K, Crooks J, et al. A conformational immunogen on VP2 of infectious bursal disease virus that induces virus-neutralizing antibodies that passively protect chickens [J]. J Gen Virol,1989,70(6):1473-1481.
[7]Heine H G, Haritou M, Failla P, et al. Sequence analysis and expression of the host-protective immunogen VP2 of a variant strain of infectious bursal disease virus which can circumvent vaccination with standard type I strains [J]. J Gen Virol,1991,72(8):1835-1843.
[8]曹冰玉,王永山,范红结,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因在大肠杆菌中的表达[J].中国兽医科学,2010,40(2)：130-134.
[9]Ulrich R, Nassal M, Meisel H, et al. Core particles of hepatitis B virus as carrier for foreign epitopes [J].Adv Virus Res,1998,50:141-182.
[10]Prel A, Le Gall-Recule G, Jestin V. Achievement of avian influenza virus-like particles that could be used as a subunit vaccine against low-pathogenic avian influenza strains in ducks [J]. Avian Pathol,2008,37 (5):513-520.
[11]Cui X, Nagesha H S, Holmes I H. Mapping of conformational epitopes on capsid protein VP2 of infectious bursal disease virus by fd-tet phage display [J]. J Virol Methods,2003,114 (1):109-112.
[1]卡尔尼克B w主编.禽病学[M].第10版.高福,苏敬良,主译.北京：中国农业出版社,1999：914-937.
[2]Muller H, Islam M R, Raue R. Research on infectious bursal disease—the past, the present and the future [J]. Vet Microbiol,2003,97 (1-2):153-165.
[3]Kibenge F S, Jackwood D J, Mercado C C. Nucleotide sequence analysis of genome segment A of infectious bursal disease virus [J]. J Gen Virol,1990,71 (3):569-577.
[4]Yamaguchi T, Lwata K, Kobayashi M, et al. Epitope mapping of capsid proteins VP2 and VP3 of infectious bursal disease virus [J]. Arch Virol,1996,141 (8):1493-1507.
[5]Heine H G, Hariton M, Failla P, et al. Sequence analysis and expression of the host-protective immunogen VP2 of a variant strain of infectious bursal disease virus which can circumvent vaccination with standard type I strains [J]. J Gen Virol,1991,72 (8):1835-1843.
[6]Cui X, Nagesha H S, Holmes I H. Mapping of conformational epitopes on capsid protein VP2 of infectious bursal disease virus by fd-tet phage display [J]. J Virol Methods,2003,114 (1):109-112.
[7]Yao K, Vakharia V N. Induction of apoptosis in vitro by the 17-kDa nonstructural protein of infectious bursal disease virus:possible role in viral pathogenesis [J]. Virology,2001,285 (1):50-58.
[8]王永山,范红结,李银,等.传染性法氏囊病病毒五个抗原表位短肽的鉴定与序列分析[J].中国病毒学,2005,20(5)：503-506.
[9]祁小乐,王笑梅,高玉龙,等.鸡传染性法氏囊病病毒VP2蛋白研究进展[J].中国预防兽医学报,2008,30(8)：656-660.
[10]王永山,欧阳伟,潘群兴,等.近期引起免疫失败的传染性法氏囊病病毒VP2基因的分子特征[J].中国兽医科学,2008,38(12)：919-925.
[11]曹冰玉,王永山,范红结,等.引起免疫失败的传染性腔上囊病病毒VP2基因在大肠杆菌中的表达[J].中国兽医科学,2010,40(2)：130-134.
[12]王军,李银,范红结,等.传染性法氏囊病病毒抗原表位分析一单克隆抗体的制备与鉴定[J].中国预防兽医学报,2005,27(3)：171-174.
[13]王永山,郝崇,侯世宽,等.抗鸡IgG单克隆抗体的研究[J].兽医大学学报,1989,9(2)：109-114.
[14]余抒,罗萍,陈洪章,等.抗肠出血性大肠杆菌O157:H7 EspA单克隆抗体的制备及其特异性鉴定[J].细胞与分子免疫学杂志,2007,23(7)：657-659.
[15]刘玉斌,苟仕金,主编.动物免疫学实验技术[M].吉林：吉林科学技术出版社,1989：150-163.
[16]Wang X N, Zhang G P, Zhou J Y, et al. Identification of neutralizing epitopes on the VP2 protein of infectious bursal disease virus by phage-Displayed heptapeptide library screening and synthetic peptide [J]. Viral Immunology,2005,18 (3):549-557.
[17]王忠灿,王永山,唐雨德,等.传染性法氏囊病病毒检测系统的建立[J].中国兽医学报,2008,28(9)：1015-1019.
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