MTT法测定大肠杆菌活菌数实验研究

Experimental study on the application of MTT colorimetric method in counting live Escherichia coli

实验分析MTT比色法中的工作波长、反应时间、MTT用量、菌龄、二甲基亚砜(DMSO)等因素对大肠杆菌活细菌计数的影响.同时,通过与平板计数对比,确定了MTT细菌计数的可行性.结果表明,当细菌数在107～109cell·mL-1范围内时,双波长MTT法可用于检测大肠杆菌的活菌数量,最佳反应时间为60min,MTT溶液用量200μL,溶解剂DMSO用量1mL,且该方法对菌龄不敏感.OD510-690值与细菌浓度C呈良好的正向线性关系,其可决系数在0.99以上.

Experimental studies were conducted on the influences of working wavelength,reaction range of microbial cell number,reaction time,concentration of MTT,cell age of Escherichia coli（E.coli）,DMSO in the MTT colorimetric method（MTT method）,etc.for determining live E.coli.Compared with dilution-plate method,the MTT method was a viable method for counting live E.coli,provided the number of bacteria was in the range 107 cell · mL-1~109 cell · mL-1.The best working parameters were reaction time 60 minutes,MTT 200 μL and DMSO 1 mL.The method was insensitive to cell age of E.coli.The correlation between the values of OD510-690 and the microbial cell number was above 0.99.