人体外周血CD3^+CD4^-CD8^-双阴性T细胞体外扩增的实验研究

Amplification of Human peripheral blood CD3^+CD4^-CD8^- double negative T cells in vitro

目的探讨人体外周血CD3^+CD4^-CD8^-双阴性T细胞(DNT细胞)在体外进行分离和扩增的方法。方法取健康的成人外周血20ml,应用Rosettesep抗体吸附法去除CD4^+T细胞和CD8^+T细胞;再将DNT细胞放入anti-CD3mAb包被的培养板中,并加入rhIL-2、rhIL-4,共同培养,第10天和第14天计数细胞扩增倍数及记录扩增曲线;利用Easysep免疫磁珠法纯化,采用流式细胞仪检测其纯度。结果经分选、纯化后的DNT细胞,纯度可达94%以上;体外培养第10天和第14天,DNT细胞扩增倍数分别为53倍和41倍。结论通过Rosettesep抗体吸附法及Easysep免疫磁珠法分离及纯化DNT细胞是可行的,可获得大量高纯度的DNT细胞。

Objective To detect the method of isolation and amplification CD3^＋CD4^-CD8^- double negative T cells（DNT）in vitro.Methods We used the Rosettesep to remove CD4^＋T cells and CD8^＋T cells from human peripheral blood.To culture DNT cells with anti-CD3 mAb,rhIL-2and rhIL-4.We computed and recordedthe amplification times after day 10 and day 14.To putify DNT cells with Easysep and test its purity with flow cytometry.Results The purity of DNT cells is 94%.The amplification times is 53 in day 10 and 41in day 14.Conclusion The method of Rosettesep and Easysep can be used to isolate and purify DNT cells.