| 肖国凯,郝珮羽,冯世涛,等.兔抗条纹斑竹鲨IgNAR多克隆抗体的制备和应用[J].中国海洋药物,2022,41(2):37-42. |
| 兔抗条纹斑竹鲨IgNAR多克隆抗体的制备和应用 |
| Preparation and application of rabbit polyclonal antibody against Immunoglobulin new antigen receptor of whitespotted bamboo shark |
| 投稿时间：2021-04-22 修订日期：2021-05-20 |
| DOI： |
| 中文关键词: 条纹斑竹鲨、IgNAR、单域抗体、多克隆抗体 |
| English Keywords:Whitespotted bamboo shark IgNAR Single domain antibody Polyclonal antibody |
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| 中文摘要: |
| 目的 制备条纹斑竹鲨IgNAR的多克隆抗体,为条纹斑竹鲨单域抗体的开发提供工具。方法 合成条纹斑竹鲨IgNAR恒定区的一段多肽作为抗原,免疫新西兰大白兔,制备多克隆抗体血清,采用间接ELISA法测定抗体的效价,并通过Western blotting检测多克隆抗体的特异性。此外,利用制备的多克隆抗体对条纹斑竹鲨外周血淋巴细胞进行免疫荧光和流式分析。结果 制备的抗血清效价高达1000万,该多克隆抗体不仅能识别条纹斑竹鲨的IgNAR,还能识别护士鲨的IgNAR,但对条纹斑竹鲨的灵敏度更高。通过免疫荧光和流式细胞术确定了该多克隆抗体还可以识别条纹斑竹鲨外周血淋巴细胞跨膜型IgNAR。结论 成功制备了条纹斑竹鲨IgNAR的多克隆抗体,该抗体可用于Western blotting、免疫荧光和流式细胞术等实验,为条纹斑竹鲨单域抗体的开发提供了有力工具。 |
| English Summary: |
| Objective This study was aimed to prepare a rabbit polyclonal antibody against the Immunoglobulin new antigen receptor (IgNAR) of whitespotted bamboo shark, and provided a tool for the development of single domain antibody of whitespotted bamboo shark. Methods We synthesized a peptide with the sequence from whitespotted bamboo sharks IgNAR constant regions and immunized New Zealand white rabbits. The polyclonal antibody was prepared, and the titer of the antibody was determined by indirect ELISA. The specificity of the polyclonal antibody to IgNAR was determined by Western blotting. In addition, the polyclonal antibody was used to detect the transmembrane IgNAR of whitespotted bamboo shark peripheral blood lymphocytes by immunofluorescence and flow cytometry. Results The titer of the antiserum was 10000000. The polyclonal antibody could detect IgNAR from both whitespotted bamboo shark and nurse shark through Western blotting, and had a higher sensitivity to the IgNAR of whitespotted bamboo shark. The polyclonal antibody could recognize transmembrane IgNAR of whitespotted bamboo shark peripheral blood lymphocytes by immunofluorescence and flow cytometry. Conclusion The polyclonal antibody against IgNAR of whitespotted bamboo shark was successfully produced. It can be used in Western blotting, immunofluorescence and flow cytometry, which will be a useful tool for the development of single domain antibody from whitespotted bamboo shark. |
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