| 猪腺病毒3型Hexon蛋白多克隆抗体制备与免疫活性鉴定 |
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| 引用本文： | 汪镇南,唐青海,颜爱,刘云福. 猪腺病毒3型Hexon蛋白多克隆抗体制备与免疫活性鉴定[J]. 中国畜牧兽医, 2015, 42(1): 1-8. DOI: 10.16431/j.cnki.1671-7236.2015.01.001 |
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| 作者姓名： | 汪镇南 唐青海 颜爱 刘云福 |
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| 作者单位： | 1. 中国农业大学动物医学院, 北京 100193;2. 湖南九鼎集团兽药公司兽医中心, 岳阳 414000;3. 南阳师范学院南阳市兽医生物工程技术研究中心, 南阳 473061 |
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| 基金项目： | 国家自然基金青年基金(31101837);河南省重点科技攻关项目(142102110101);南阳师范学院引进人才专项项目(70640) |
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| 摘 要： | 本研究旨在制备猪腺病毒3型(PADV-3)Hexon蛋白多克隆抗体.试验构建重组原核表达载体pET28a-Hexon,IPTG诱导重组蛋白的表达并进行Western blotting鉴定,目的蛋白与佐剂混合、乳化制备免疫原,免疫家兔制备Hexon蛋白多克隆抗体.采用免疫过氧化物酶单层细胞染色法(IPMA)检测抗体的免疫活性与抗体滴度.结果显示,Hexon蛋白原核表达以包涵体形式存在,分子质量约为105 ku;真核细胞表达的Hexon蛋白均定位于HEK293细胞质内,细胞核内无分布;IPMA测定所制备的多克隆抗体滴度为1:1 600,该抗体与体外培养的PADV-3及稳定表达Hexon蛋白的细胞均呈特异性反应.结果表明本试验制备的PADV-3型Hexon蛋白多克隆抗体免疫活性和特异性良好.
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| 关 键 词： | 猪腺病毒3型 Hexon基因 多克隆抗体 |
| 收稿时间： | 2014-08-02 |
Preparation and Immuno-activity Identification of Polyclonal Antibodies against Porcine Adenovirus Serotype 3 Hexon Protein |
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| WANG Zhen-nan,TANG Qing-hai,YAN Ai,LIU Yun-fu. Preparation and Immuno-activity Identification of Polyclonal Antibodies against Porcine Adenovirus Serotype 3 Hexon Protein[J]. China Animal Husbandry & Veterinary Medicine, 2015, 42(1): 1-8. DOI: 10.16431/j.cnki.1671-7236.2015.01.001 |
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| Authors: | WANG Zhen-nan TANG Qing-hai YAN Ai LIU Yun-fu |
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| Affiliation: | 1. College of Veterinary Medicine, China Agricultural University, Beijing 100193, China;2. Center for Veterinary Medicine, Hunan Jiuding Group Veterinary Pharmaceutical Company, Yueyang 414000, China;3. Center for Nanyang Veterinary Biological Engineering Technology, Nanyang Normal University, Nanyang 473061, China |
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| Abstract: | This study was aimed to prepare polyclonal antibodies against porcine adenovirus serotype 3 (PADV-3) Hexon protein.The recombinant prokaryotic expression plasmid pET28a-Hexon was constructed and transformed into BL21 (DE3), 1 mmol/L concentration of IPTG was used to induce the target gene expression which was then analyzed by SDS-PAGE and Western blotting.The expressed proteins were emulsified with freund''s adjuvant, the mixture was used to immunize the rabbit to prepare the polyclonal antibodies against PADV-3 Hexon protein.The immunocompetence and titers of the antibody were identified by immunoperoxidase monolayer assay (IPMA).The results showed that prokaryotic expressed Hexon protein existed as inclusion bodies, with a molecular weight of 105 ku, eukaryotic expressed Hexon protein was only observed in the cytoplasm of HEK293 cells.PADV-3 Hexon-specific polyclonal antibody showed perfect activity which recognized the PADV-3 propagated in PK-15 cells and the recombinant Hexon protein expressed in HEK293 cells specifically, it was IPMA antibody positive when diluted in 1:1 600 ratio.The prepared PADV-3 Hexon polyclonal antibodies showed wonderful activity and specificity. |
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| Keywords: | porcine adenovirus serotype 3 Hexon gene polyclonal antibody |
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