Biofilm imaging by scanning electron microscopy

Glass coverslips were placed in 6-well plates, bacterial suspension was added to the wells and incubated at 37°C in a rotary incubator for 24 hours to facilitate biofilm formation. The bacteria were treated with propolis Eth or EA extracts or CPNP. Untreated control was included. After incubation, the glass coverslips were rinsed with saline followed by fixation with 2.5% glutaraldehyde. Biofilms were serially dehydrated, air-dried and sputter coated with gold (SC7620 Mini Sputter Coater, Quorum Technologies, UK). The biofilms formed were observed using a TM3000 tabletop scanning electron microscope (Hitachi, Japan). The experiments were carried out in triplicates with three independent repeats.