Wang X M, Terasaki P I, Rankin G W, Chia D, Zhong H P, Hardy S
UCLA Tissue Typing Laboratory, Department of Surgery, UCLA School of Medicine 90024.
Hum Immunol. 1993 Aug;37(4):264-70. doi: 10.1016/0198-8859(93)90510-8.
We present a microtest for cell-mediated immunity, based on the use of the Tarasaki tray and calcein AM vital dye. The number of target cells needed has been reduced to 500 per test with a corresponding tenfold reduction in the number of effector cells needed. Results were read at the rate of 1 second per test using a fluorimeter attached to a microscope. Each reaction was also confirmed visually with the use of ethidium bromide as a counterstain for dead cells. The calcein AM dye used to stain the living cells was shown to have a low spontaneous leakage rate--less than 15% in 4 hours at 37 degrees C. Dilutions of targets stained by calcein AM had a linear relationship with measured fluorescence values. NK cells, LAKs, and CTLs were readily detectable by this microtest. Quantitation of killing and kinetic analysis was readily performed with this test system. A significant positive correlation to 51Cr-release results was found. We conclude that the microtest should find wide application in studies of cell-mediated immunity.
我们展示了一种基于使用田崎托盘和钙黄绿素 AM 活细胞染料的细胞介导免疫微试验。每次试验所需的靶细胞数量已减少至 500 个，相应地所需效应细胞数量减少了十倍。使用连接到显微镜的荧光计，以每次试验 1 秒的速度读取结果。每次反应还用溴化乙锭作为死细胞的复染剂进行了肉眼确认。用于染色活细胞的钙黄绿素 AM 染料显示出自发泄漏率低——在 37 摄氏度下 4 小时内低于 15%。经钙黄绿素 AM 染色的靶细胞稀释液与测量的荧光值呈线性关系。通过这种微试验很容易检测到自然杀伤细胞、淋巴因子激活的杀伤细胞和细胞毒性 T 淋巴细胞。使用该测试系统可以很容易地进行杀伤定量和动力学分析。发现与 51Cr 释放结果有显著正相关。我们得出结论，该微试验应在细胞介导免疫研究中得到广泛应用。
