Figure 3

Bacteriophage-based E. coli detection assay workflow. (a) Sterile tap water samples were spiked with varying concentrations of E. coli and incubated shaking at 37 °C for 3 h to resuscitate cultures. (b) Bacteriophage immobilized to magnetic nanoparticles were added to the sample and incubated shaking for 10 min at 37 °C. (c) E. coli cells in the sample were captured by the bacteriophage immobilized nanoparticles. (d) A magnet was used to concentrate the bacteria-bacteriophage nanoparticle complex allowing the supernatant removal. (e) After 3 h of incubation (shaking at 37 °C) to allow for luciferase reporter enzyme expression and bacteria lysis to occur, samples were filtered through a 96 well filter plate to capture the luciferase reporter enzyme. (f) Luciferase substrate was added to each well and incubated at room temperature for 10 min for reaction to occur. (g) Luminescent signal was read in a spectrophotometer.