Abstract The present article reports on the influence of various atrazine concentrations to the response of genetically modified Escherichia coli TV1061 bacterial cells while modulating the experimental conditions. Interesting increases of bioluminescence signals are recorded for E. coli TV1061 bacteria in the presence of 10μg/mL atrazine concentration named “high-toxicity bacteria alert” when compared with 1μg/mL –10fg/mL atrazine termed “low-toxicity bacteria alert”. Detecting the effect of atrazine via its effect on bioluminescence of bacteria has been carried out by two consecutive measurements (fresh and overnight modes) at different concentrations of analyte. We have shown that a more precise discrimination at lower-toxicity concentrations can be obtained through overnight incubation of bacteria with the analyte at 4°C. In addition, centrifugation of bacterial cells and analyte dilutions has been performed in order to ensure a better interaction between the insoluble atrazine pesticide and the bacterial cells.