免疫磁珠兩步法分離大鼠脾臟CD4+CD25+調節性T細胞_《中國普外基礎與臨床雜志》

作者：

浦立勇 , 王學浩 , 李相成 , 張峰 , 俞悅 , 姚愛華 , 成峰 , 李國強

南京醫科大學第一附屬醫院肝臟移植中心（南京210029）;;
通訊作者：王學浩;

關鍵詞：

磁性細胞分離 CD4+CD25+調節性T細胞流式細胞儀技術大鼠

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目的探索利用磁性細胞分離(MACS)系統高效、快速地分離大鼠脾臟CD4+CD25+調節性T細胞。
方法采用免疫磁珠兩步法分離大鼠脾臟內的CD4+CD25+調節性T細胞, 首先采用“雞尾酒”抗體和抗IgG磁珠陰性分選CD4+ T細胞，再用抗CD25PE抗體和抗PE磁珠陽性分選獲得CD4+CD25+T 細胞。分離后的細胞經流式細胞儀檢測分離純度；臺盼藍染色檢測細胞存活率；體外增殖實驗檢測其對CD4+CD25-T細胞的免疫抑制作用。
結果陰性分選獲得的CD4+T細胞純度為（83.6±2.5）%（79%~87%）, 陽性分選后獲得的CD4+CD25+T細胞純度為（90.2±1.8）%（86%~93%）, 細胞存活率為（92.8±3.4）%（92%~95%）, 體外增殖實驗表明，CD4+CD25+T細胞能明顯抑制CD4+CD25-T 細胞的增殖（P＜0.01）。
結論采用MACS系統陰性加陽性分選可以高效、快速地獲得理想純度和有免疫抑制功能的大鼠CD4+CD25+調節性T細胞。

引用本文： 浦立勇,王學浩,李相成,張峰,俞悅,姚愛華,成峰,李國強. 免疫磁珠兩步法分離大鼠脾臟CD4+CD25+調節性T細胞. 中國普外基礎與臨床雜志, 2006, 13(3): 275-278. doi: 復制

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5. [Ｊ]. Blood, 2002; 99(10)∶3493.
6. [Ｊ]. Blood, 2003; 102(6)∶2180.
7. [Ｊ]. J Exp Med, 2001; 193(11)∶1303.
8. [Ｊ]. Blood, 2004; 104(3)∶895.
9. Stephens LA, Barclay AN, Mason D. Phenotypic characterization of regulatory CD4+CD25+ T cells in rats.
10. Sakaguchi S, Sakaguchi N, Asano M, et al. Immunologic selftolerance maintained by activated T cells expressing IL2 receptor alphachains (CD25). Breakdown of a single mechanism of selftolerance causes various autoimmune diseases.
11. SuriPayer E, Amar AZ, Thornton AM, et al. CD4+CD25+ T cells inhibit both the induction and effector function of autoreactive T cells and represent a unique lineage of immunoregulatory cells.
12. Takahashi T, Kuniyasu Y, Toda M, et al. Immunologic selftolerance maintained by CD25+CD4+ naturally anergic and suppressive T cells: induction of autoimmune disease by breaking their anergic/suppressive state.
13. Taylor PA, Lees CJ, Blazar BR. The infusion of ex vivo activated and expanded CD4+CD25+ immune regulatory cells inhibits graftversushost disease lethality.
14. Jiang S, Camara N, Lombardi G, et al. Induction of allopeptidespecific human CD4+CD25+ regulatory T cells ex vivo.
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16. Hoffmann P, Eder R, KunzSchughart LA, et al. Largescale in vitro expansion of polyclonal human CD4+CD25+ high regulatory T cells.
17. Ikemoto T, Tashiro S, Yasutomo K, et al. Donorspecific tolerance induced by simultaneous allogeneic islet transplantation with CD4+CD25+ Tcells into hepatic parenchyma in mice [Ｊ]. J Med Invest, 2004; 51(3-4)∶178.
18. 徐勇, 霍梅. 免疫磁珠分離及流式細胞儀分選純化外周血CD34+/CD90+ 干細胞 [Ｊ]. 臨床檢驗雜志, 2004; 22(4)∶246.

《中國普外基礎與臨床雜志》

免疫磁珠兩步法分離大鼠脾臟CD4+CD25+調節性T細胞

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《中國普外基礎與臨床雜志》

免疫磁珠兩步法分離大鼠脾臟CD4+CD25+調節性T細胞

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