四唑氮蓝法检测胶质瘤碱性蛋白iRNA诱导的细胞毒作用

3- (4,5-dimethylthiazol-2-yl) - 2,5-diphenyl-tetrazolium bromide colorimetric assay for cytotoxity induced by glioma basic proteinspecific immune RNA

作者以人脑胶质瘤细胞系BT_(325)作为靶细胞,采用四唑氮蓝(MTT)标记计数细胞的方法,观察了胶质瘤碱性蛋白iRNA致敏的小鼠脾淋巴细胞对肿瘤细胞的细胞毒作用,并与同位素释放法进行了比较.结果表明,在2.5×10~3～4×10~4个肿瘤细胞范围内,MTT还原产物甲(月朁)吸光度与细胞数和保温时间成正比.选靶细胞数2×10~4个和保温时间4 h作细胞毒检测,在效/靶比例分别为25:1、50:1和100:1时,MTT法所得细胞毒指数依次为11.93％,25.11％和41.12％;~3H ^-Leu法指数为11.50％,26.04％和40.65%,统计分析两法数值P>0.05,即MTT法用于细胞毒测定是可行的.

3 - (4,5-dimethylthiazol-2-yl) - 2,5-diphenyl tetrazolium bromide (MTT) colorimetric assay was employed in a cytotoxity test, in which a human glioma cell line, BT32.5 was used as target cells, and mouse splenocytcs activated by glioma basic protein-specific iRNA (GBP-iRNA) as effector cells. A comparison of the cytotoxily obtained by the method with that by 3H-Leu cytotoxity assay was performed. The results show that the amount of MTT reduced to formazane is proportional to the cell numbers and incubation lime within the range of 2.5×103 -4 ×104 cells for 3-5 h; and 2 × 104 cells for 4 h, that the cytotoxity of MTT assay at 25 : 1, 50 : 1 and 100 : 1 ratio of effector cells to target cells is 11.93%, 25.11% and 41.12% respectively; and that the counterpart of 3H Leu assay is 11.50%, 26.04% and 40.65%. Statistical analysis of the respective values showed P>0.05, hence the 3H-Leu assay could be replaced by MTT-assay.