- 中文核心期刊
- CSCD来源期刊
- 中国科技核心期刊
- CA、CABI、ZR收录期刊
| Citation: |
ZHU D W, XU Y Q, YIN Y Y, et al. Preparation of Monoclonal Antibodies Against CD25 of Porcine Tregs [J]. Fujian Journal of Agricultural Sciences，2024，39（12）：1336−1345. DOI: 10.19303/j.issn.1008-0384.2024.12.003
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The monoclonal antibody against CD25 in pigs was obtained to facilitate detecting and studying the regulatory T cells (Tregs).
The porcine 6×His tagged CD25 fragment was cloned into prokaryotic expression vector pET-28a via homologous recombination to construct the recombinant plasmid pET28a-CD25. Subsequently, the 6×His-CD25 recombinant protein was expressed in E. coli and purified on a metal chelating affinity chromatography medium (Ni-NTA) pre-packed column to be used as the antigen to immunize female BALB/c mice. Spleen cells of the immunized mice were hybridized with myeloma cells SP2/0 and screened by indirect ELISA prior to culture cloning. The target monoclonal antibody against CD25 was thus prepared using mouse ascites method.
A single cell-derived hybridoma cell line with a stable expression of CD25 monoclonal antibody was successful obtained. ELISA and western blotting showed the antibody had a strong specificity with an exclusive reactivity towards the target protein, not the others, even at a high titer of 1:
The recombinant porcine CD25 was successfully prepared and the specific monoclonal antibody obtained for qualitative and/or quantitative detection of the protein as well as further studies on the functions of porcine Tregs.
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