AN IMPROVED COLORIMETRIC ASSAY FOR CELL-PROLIFERATION AND VIABILITY UTILIZING THE TETRAZOLIUM SALT XTT
AN IMPROVED COLORIMETRIC ASSAY FOR CELL-PROLIFERATION AND VIABILITY UTILIZING THE TETRAZOLIUM SALT XTT
复制标题
DOI：
10.1016/0022-1759(91)90114-u
复制
发表时间：
1991-09-13
影响因子：
2.2
通讯作者：
GLASEBROOK, AL
中科院分区：
文献类型：
--
作者：
ROEHM, NW;RODGERS, GH;GLASEBROOK, AL
一种新的四氮唑盐XTT，3‘-[1-[(phenylamino)-carbonyl]-3，4-tetrazolium]-bis(4-methoxy-6-nitro)benzene-sulfonic酸水合物，被用于正常激活的T细胞和几种细胞因子依赖的细胞株的细胞活力和增殖的比色分析。XTT被代谢活性细胞的脱氢酶裂解，产生一种高度有色的、水溶性的福马赞产物。这一特点消除了在吸光度测量之前需要福马赞晶体增溶的需要，当使用其他四唑盐时，如使用四甲基偶氮唑盐时。所有被研究的小鼠细胞对XTT的生物还原并不是特别有效，但可以通过添加电子偶联剂如吩嗪甲硫酸盐(PMS)或甲萘二酮(MEN)来增强。用T细胞系HT-2和11.6、NFS60a、肥大细胞系MC/9a和有丝分裂原激活的脾T细胞，确定了PMS或MEN对XTT代谢的最适浓度。当与经前综合征联合使用时，每个细胞在XTT作用下产生的Formazan吸光度值都比用四甲基偶氮唑盐观察到的高。因此，XTT在比色细胞增殖分析中的使用具有明显的优势，因为它减少了分析时间和样品处理，同时提供了同等的灵敏度。
A new tetrazolium salt XTT, sodium 3'-[1-[(phenylamino)-carbonyl]-3,4-tetrazolium]-bis(4-methoxy-6-nitro)benzene-sulfonic acid hydrate, was evaluated for use in a colorimetric assay for cell viability and proliferation by normal activated T cells and several cytokine dependent cell lines. Cleavage of XTT by dehydrogenase enzymes of metabolically active cells yields a highly colored formazan product which is water soluble. This feature obviates the need for formazan crystal solubilization prior to absorbance measurements, as required when using other tetrazolium salts such as MTT. Bioreduction of XTT by all the murine cells examined was not particularly efficient, but could be potentiated by addition of electron coupling agents such as phenazine methosulfate (PMS) or menadione (MEN). Optimal concentrations of PMS or MEN were determined for the metabolism of XTT by the T cell lines HT-2 and 11.6, NFS-60 a myeloid leukemia, MC/9 a mast cell line and mitogen activated splenic T cells. When used in combination with PMS, each of these cells generated higher formazan absorbance values with XTT than were observed with MTT. Thus the use of XTT in colorimetric proliferation assays offer significant advantages over MTT, resulting from reduced assay time and sample handling, while offering equivalent sensitivity.