Preparation of biofilm assay using 96-well and 6-well microplates for quantitative measurement and structural characterization: a review

The term "microbial biofilm" refers to three-dimensional, stationary populations of microorganisms comprising various cell types encased in a matrix primarily produced by the microbial community. In biofilm research, selecting the appropriate experimental platform is crucial, as it determines the data gathered and, consequently, impacts the quality of the studies. Biofilm biomass and viability can be assessed using the crystal violet assay and resazurin assay, respectively. Due to its convenience and adaptability, the 96-well microplate is frequently employed in various scientific and medical applications. Various characterization techniques for biofilm assays using the 6-well microplate include light microscopy, confocal laser scanning microscopy (CLSM), scanning electron microscopy (SEM), atomic force microscopy (AFM), and FTIR (Fourier-transform infrared) spectroscopy. This mini review discusses the measurement of biofilm biomass and viability, experimental protocols for crystal violet and resazurin assays, and potential issues in microplate-based biofilm assays.