MTT法检测细菌细胞数的主要影响因素分析

Analysis of Influencing Factors on the Bacterial Cell Counting with the MTT Method

MTT法是生物活细胞计数的灵敏、快速和便捷方法。报告对MTT法应用于细菌细胞数定量分析时的分析前培养时间、MTT反应时间、MTT剂量和检测波长等主要影响因素的分析结果,同时报告MTT反应产物溶解检测与直接检测,以及MTT法与平板菌落计数法定量检测的结果比较。结果如下:Formazan生成量与细菌活细胞数呈正相关,对数生长期变化最明显;MTT为0.25 mg/mL,反应2 h时吸光值与菌落数具有良好的线性关系(r=0.999 8);Formazan的最大吸收波长为570～580 nm;MTT反应产物溶解检测和直接检测结果经统计学分析无显著性差异(P>0.05);细菌细胞数在107～109 cfu/mL范围内,MTT法与菌落计数法定量结果呈现良好的线性关系(r≥0.991 7)。

MTT method is a sensitive,rapid,and simple cytometry for counting living bio-cell.This paper reported the results of analysis when applying MTT method on bacterial cell quantity analysis,incubation time before the analysis,MTT reaction time,MTT dosage and detection wavelength and other main effective factors.At the same time also reported the comparison results of the lysing detection of reaction product with direct detection of it,and MTT method with plate colony counting method.The results were as follows：（1） the production of Formazan assumed positive pertinence to the quantity of living bacterial cells,the change assumed the most obvious at logarithmic phase;（2） Absorbancy value possessed fine linear relation（r = 0.999 8） with colony quantity when the reaction time was 2 hours with 0.25 mg/mL MTT;（3） The maximum absorption wavelength was 570～580 nm;（4） The statistical results showed that there was no significant difference in the reaction products between lysing detection of reaction product and direct detection（P0.05）;（5） bacterial cell quantity within the range of 107～109 cfu/mL,the result assumed good linear relationship（r ≥ 0.991 7） between the MTT method and the plate colony counting method.