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Crystal violet biofilm assays

CB Connor J. Beebout
LS Levy A. Sominsky
AE Allison R. Eberly
GH Gerald T. Van Horn
MH Maria Hadjifrangiskou
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Determination of biofilm biomass was performed using the crystal violet assay as previously described33. Overnight cultures were diluted to optical density 600 nm (OD600) = 0.05, and 100 µL of the diluted culture was aliquoted into a 96-well polyvinyl chloride (PVC) plate (Costar). Plates were incubated in a humid chamber at room temperature, washed and stained with crystal violet, and disaggregated using 35 percent acetic acid. For antibiotic assays, biofilms were grown for 48 h, antibiotics or vehicle was added to each well, and biofilms were grown another 72 h prior to determining biomass. Total biomass was determined by measuring the absorbance at 570 nm using a SpectraMax i3 microplate reader (Molecular Devices).

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