鳜免疫球蛋白单克隆抗体的制备及特性

Production and characterization of monoclonal antibodies against Siniperca chuatsi Ig

  • 摘要: 鳜(Siniperca chuatsi)是我国传统名贵淡水鱼。近年来,随着鳜养殖业的发展,鳜病害也日益猖獗。暴发性疾病的发生与流行阻碍了鳜养殖业的健康发展。要解决这些问题,除了加强管理、提高养殖技术外,还必须重视其病原学、免疫学等领域的研究,将免疫预防手段引入到鳜养殖业,通过免疫手段有效增强鱼体主动防御能力,减少疾病发生机率。然而对鳜免疫学的研究还处于初始状态,张永安等提纯了鳜血清免疫球蛋白并进行亚单位分子量的测定。

     

    Abstract: This paper described the preparation and characterization of monoclonal antibodies (Mab) against mandarin fish( Sinipewa chuatsi) immunoglobulin(Ig), using the Mabs as a tool for analyzing the structure of S. chuatsi Ig, and monitoring the humoral immunity level of S. chuatsi. The purified serum Ig of S. chuatsi was used as antigen to immunize Balb/C mice, after cell fusion and screening the antibody secreting cell, eleven hybridoma cell lines which secreting Mab against S. chuatsi Ig had been established. The characteristics of these Mabs had been analyzed. In isotyping analysis, the results showed that two of them were IgM, three of them were IgG3, five of them were IgG1 and one of them belonged to IgA; Further experiments proved that Mab 7F12-F6 only specifically bound to the Ig of S. chuatsi; MablF12-A3,4D3-A7,3F8-H3,3A5-D11,5C3-A7,5D11-F9 and 6G6-All were able to recognise the Ig of S. chuatsi, Anguilla anguilla and Auguilla japonica. And Mab3D5-H7,3D3-A6,4Cl-H11 had the cross reaction with the Ig of A. anguilla, A. japonica and Colossoma brachypomum. All of Mabs did not have any cross reaction on the Ig of Pseudosciaena crocea, Epinephelus drummondhayi, Carrassius auratus, Hybrid tilapia, Clarias fuscus, ornamental carp, Micropterus salmonoides and the fish common bacterial pathogens, such as Aeromonas, Edwardsiella, Vibrio, Salmonella and Escherichia coli. And their ELISA titer ranged from 104 to 106, the sensitivity of the Mabs purified Siniperca chuatsi Ig was at least lower than 39 ng? mL^ - 1. These results suggested that the different fish species had both species specific epitopes and the common epitopes on their Ig. These common epitopes not only existed between the close relative species, such as the Japanese eel and European eel, but also existed between the distant relative ones such as S. chuatsi, eel and C. brachypomum cuvier. The studies revealed that the Ig of S. chuatsi had evolutionary relationship with the Ig of eel and C. brachypomum cuvier, and the specific Mab has the potential to be used for detecting the humoral immunity level and the pathogen of S. chuatsi.

     

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