Live dead baclight bacteria viability stains

CLSM was used to confirm the effect of peptide 8 on mature biofilm respect the controls. MSRP were grown in chambered cover glass (μ Slide 4 well; ibidi GmbH, Germany) in a static condition for 24 h. Peptide 8 was added on a 1-day-old biofilm at 12.5 μM. Bacterial suspensions incubated with medium alone were used as a positive control. After 24 h, biofilms were rinsed with PBS and stained by using a LIVE/DEAD^® BacLight Bacteria Viability stains (Life Technologies, Italy). After the staining, the images were observed using a LSM 700 inverted confocal laser-scanning microscope (Zeiss, Italy). The areas were scanned using a 10X objective lens with the signal recorded in the green channel for Syto9 (excitation 488 nm, emission 500–525 nm) and in red channel for PI (excitation 500–550 nm, emission 610–650 nm).

Confocal laser scanning microscopy (CLSM) was used to illustrate the effect of abietic acid on viability and architecture of mature biofilms of MSSP and MRSP1. Cells were grown in chambered cover glass (μ Slide 4 well; ibidi GmbH, Gräfelfing, Germany) in a static condition. Abietic acid was added on a 1-day-old biofilm at 2 × MIC value for each strain. After 24 h, biofilms were rinsed and stained by using a LIVE/DEAD^® BacLight Bacteria Viability stains (Life Technologies, Monza, Italy). The images were observed using an LSM 700 inverted confocal laser-scanning microscope (Zeiss, Arese, Milano, Italy), using a 10× objective lens with the signal recorded in the green channel for Syto9 (excitation 488 nm, emission 500–525 nm) and in red channel for PI (excitation 500–550 nm, emission 610–650 nm).