Cedillo-Rivera R, Ramírez A, Muñoz O
Unidad de Investigación Clínica en Enfermedades Infecciosas y Parasitarias, Hospital de Pediatría, Centro Médico Nacional Siglo XXI, IMSS, México, DF.
Arch Med Res. 1992;23(2):59-61.
A rapid colorimetric assay for viability of E. histolytica trophozoites using the tetrazolium salt MTT and phenazine methosulfate (PMS) was developed. The MTT is normally reduced by dehydrogenases of viable cells and transformed to formazan, a colored compound; the reaction is catalyzed by PMS. Trophozoites of E. histolytica were incubated in the presence of MTT and PMS during 15, 30, 45, 60 and 75 min, at 37 degrees C. Trophozoites killed by heat or cold were used as controls. The formazan produced into the cells was extracted with isopropanol-HCl and the optical density (OD) was measured at 570 nm. There was a linear relation between the number of viable trophozoites and the OD570 (r2 = 0.98). The trophozoites killed by cold or heat did not reduce MTT. The optimal time of incubation with MTT/PMS was 45 min. The MTT/PMS assay for determining viability of E. histolytica trophozoites is reproducible, rapid and easy to perform and it could be useful to study cytotoxicity and susceptibility to drugs.
开发了一种使用四唑盐MTT和吩嗪硫酸甲酯（PMS）快速比色测定溶组织内阿米巴滋养体活力的方法。MTT通常由活细胞的脱氢酶还原并转化为甲臜，一种有色化合物；该反应由PMS催化。在37℃下，将溶组织内阿米巴滋养体在MTT和PMS存在的情况下孵育15、30、45、60和75分钟。用热或冷杀死的滋养体用作对照。用异丙醇 - 盐酸提取细胞内产生的甲臜，并在570nm处测量光密度（OD）。活滋养体数量与OD570之间存在线性关系（r2 = 0.98）。冷或热杀死的滋养体不会还原MTT。MTT/PMS孵育的最佳时间为45分钟。用于测定溶组织内阿米巴滋养体活力的MTT/PMS测定方法具有可重复性、快速且易于操作，并且对于研究细胞毒性和药物敏感性可能有用。
