硫酸脑苷酯负载的CD1d四聚体检测小鼠Ⅱ型NKT细胞

Sulfatide-loaded CD1d tetramer to detect type Ⅱ NKT cells in mice

目的:建立一种检测小鼠Ⅱ型NKT细胞的方法。方法:将硫酸脑苷酯(sulfatide)与生物素连接的CD1d单体按摩尔比3∶1混合室温静置过夜,之后加入PE标记的链酶亲和素,室温孵育4 h制成sulfatide/CD1d四聚体。然后应用流式细胞术(FCM)检测正常小鼠脾脏及肺脏单个核细胞中Ⅱ型NKT细胞的百分比(TCR-β+sulfatide/CDld四聚体+),以及小鼠脾细胞经sulfatide刺激后单个核细胞中的Ⅱ型NKT细胞的百分比。结果:正常小鼠脾脏及肺脏单个核细胞中Ⅱ型NKT细胞的比例分别为(0.875±0.096)%和(1.175±0.263)%;小鼠脾细胞经sulfatide刺激后的Ⅱ型NKT细胞的比例为(2.75±0.603)%,与正常相比明显增加(P<0.01)。结论:硫酸脑苷酯负载的CDld四聚体是检测小鼠Ⅱ型NKT细胞的有效方法。

AIM： To create a method of detecting typell natural killer T （NKT） cells of mice. METHODS： Biotinylated mouse CDld monomers were mixed with sulfatide at a molar ratio of 1 ： 3 （ protein ： lipid ） and incubated at room temperature overnight, and then 80 μg of streptavidin-PE was added into 200 tag of the CDld-sulfatide mixture and incubated at room temperature for 4 h to get sulfatide/CDld tetramer. Flow cytometry was used to detect the percentage of type H NKT cells in mononuclear cells （MNCs） of lung and spleen of normal mice, as well as the percentage of type H NKT cells in spleen MNCs of mice after stimulated with sulfatide. RESULTS： In normal mice, the percentage of type H NKT cells accounted for （0.875 ± 0.096） % and （1. 175 ±0.263） % in MNCs of spleen and lung; the percentage in spleen MNCs after activated with sulfatide was （2.75 ± 0. 603）%, which significantly increased as com- pared with that in normal mice （P〈0.01）. CONCLUSION： Sulfatide-loaded CDld tetramer is an effective method of detecting type Ⅱ NKT cells in mice.