Influence of incubation temperatures and pH values of culture liquid on growth of Escherichia coli and the fluorescent intensity of culture liquid had been investigated.E.coli incubated at 44℃ in the culture liquid with a pH value in the range of 7.0~7.5 was found to be the best condition for fluorescent assay.Single tube technology to detect E.coli was established through the studies on the relationship between incubation time,inoculation quantity of E.coli and the fluorescent intensity of culture liquid.Relatively standard diversity of single tube detection technology was lower than those of colony-counting method and the most probably number method.The results indicated that the single tube quantitative detection method was fit for E.coli quantitative detection because of the several advantages such as less time-consuming and more economical compared with traditional methods.