中国动物传染病学报2018，Vol.26Issue(1)：19-24,6.
PRRSV N蛋白抗原肽的预测、合成以及针对该抗原肽抗体的制备和应用
PREDICTION AND SYNTHESIS OF ANTIGEN PEPTIDE OF PRRSV N PROTEIN AND ITS APPLICATION FOR GENERATION OF ANTIBODY
摘要
Abstract
To date, prediction of antigen peptides based on bioinformatics, chemical synthetic and immunological techniques has been commonly used for generation of antibodies. Here, we determined the possibility of generation of specific antibody against PRRSV N protein by using predicted antigen peptide. First, one antigen peptide (16 amino acids) close to N terminal of was predicted from N protein of a highly pathogenic PRRSV strain. Then, the BLAST analysis showed that this peptide was conserved in different North American PRRSV strains. Secondly, this antigenic peptide was synthesized by using Fmoc solid phase method and coupled to the KLH vector protein by MBS method. Subsequently, New Zealand white rabbits were immunized with the conjugated antigenic peptide emulsified with Freund's complete or incomplete adjuvant. The antiserum was collected from the immunized rabbit and antibodies were purified using IgG affinity chromatography. The purified antibodies showed good signals at dilution of over 105 in ELISA. When used at dilution of 1:2000 in Western blotting, the antibodies reacted with N protein from Marc-145 cells infected with different PRRSV strains. The dilution of 1: 200 of this antibody preparation was used in IFA for detection of PRRSV in infected cells. Collectively, the antigen peptide of the PRRSV N protein was predicted using bioinformatics software and used for generation of antibodies, which showed good reactivity in ELISA, Western blot and IFA.关键词
猪繁殖与呼吸综合征病毒/核衣壳蛋白/抗原肽/抗体Key words
Porcine reproductive and respiratory syndrome virus/nucleocapsid protein/antigen peptide/antibody分类
农业科技引用本文复制引用
相笑,马志永,邱亚峰,张彦兵,石元元,李玉明,刘珂,魏建超,邵东华,李蓓蓓,童光志..PRRSV N蛋白抗原肽的预测、合成以及针对该抗原肽抗体的制备和应用[J].中国动物传染病学报,2018,26(1):19-24,6.基金项目
中央级公益性科研院所基本科研业务费(2014JB15、2015ZL065) （2014JB15、2015ZL065）
国家自然基金(31502046) （31502046）
973计划项目(2014CB542702) （2014CB542702）
