Bacterial Membrane Staining and Analysis

Control or AEA-treated bacteria were exposed to 1 µg/mL of diamidino-2-phenylindole (DAPI) for 30 min together with 10 µg/mL Nile Red (APExBIO, Boston, MA, USA) for 30 min at 37 °C. After incubation, the samples were analyzed by flow cytometry (LRS-Fortessa flow cytometer, BD Biosciences). The 355 nm laser was used for DAPI, and the fluorescence was collected by the blue (450 nm) filter. DAPI is a blue-fluorescent probe that fluoresces brightly upon selectively binding to DNA [11 (link)]. The Nile Red was analyzed using the 561 nm yellow–green laser excitation, and data were collected using the 635 nm filter [20 (link)].