Biofilm formation
The antimicrobial properties of chitosan were assessed using a
microtitre plate assay (26). Bacteria cultured overnight were inoculated
into the wells of a 96-well microtitre plate. Chitosan at concentrations
of 10 and 100 μg/mL was applied, while control wells remained untreated.
Following a 24-hour incubation, the media was aspirated, and the wells
were rinsed thrice with sterile phosphate buffer. Then, crystal violet
solution (0.1%) was added to the wells and allowed to incubate for 15
minutes. Excess dye was removed by washing the wells, and the biofilms
were dissolved in 90% ethanol. The optical density of the dissolved
biofilms was measured at 620 nm using a microtitre plate reader.