2.11. Confocal Laser Scan Microscopy (CLSM) Analysis

The, S. aureus biofilm was grown in a confocal microscopy dish (glass-bottom dish) (Bar-Naor, Petach-Tikwa, Israel) with or without supplementation of the supernatant at the same conditions as described above. After 24 h of incubation at 37 °C, the surface-unattached cells were removed by washing the dish two times using DW. Next, the biofilm cells were stained using FilmTracer LIVE/DEAD Biofilm Viability Kit (Molecular Probes, Eugene, OR, USA) and incubated for 30 min in room temperature without exposure to light. Then, the stain was washed away and analyzed by confocal laser microscopy (CLSM) (Olympus, Hamburg, Germany). Fluorescence emission from the stained samples was measured with an SP8 CLSM (Leica) equipped with 488- and 552-nm lasers.