Biofilm Formation Assay for P. aeruginosa

Before starting antibiofilm assay, the organism P. aeruginosa MTCC 2488 was tested for its capability to form biofilm. Briefly, P. aeruginosa was grown in sterile test tubes containing TSB at 37 °C for 48 h. After the incubation time, the cultures were removed and the formed biofilm was washed three times with sterile phosphate buffer saline (PBS) and stained with 0.1% (v/v) Safranin for 10 min. The excess stain was removed by washing with sterile PBS and dried overnight at 37 °C. Safranin from stained adherent P. aeruginosa were re-dissolved in 30% (v/v) glacial acetic acid and measured absorbance at 492 nm by UV-Vis spectrophotometer (Parkin-Elmer)3 (link). The experiment was performed in triplicate.