Isolation of persister cells from an exponentially growing culture. E. coli ASV cells containing this reporter cassette were grown in LB medium to mid-exponential phase (~1 × 108 cells/ml) at 37°C with aeration and sorted using a high speed cell-sorter equipped with a standard GFP filter set. (A) Two populations were detected using forward light-scatter, one that fluoresced brightly (R3), and another that did not (R4). (B) The sorted populations were visualized by epifluorescent microscopy (bar, 5 μm). (C) Cells were sorted as described in (A-B). Once sorted both populations were treated with ofloxacin (5 μg/ml) for three hours, diluted and spotted onto LB agar plates for colony counts.