Microtiter Dish Biofilm Disruption Assay

The microtiter dish biofilm formation assay²⁴ was utilized to assess the ability of the thiazole compounds to disrupt an adherent staphylococcal biofilm, similar to what has been described elsewhere^{25 (link)}. S. epidermidis ATCC 35984 was transferred to tryptic soy broth and incubated at 37°C for 24 h before being diluted 1:200 in tryptic soy broth + 1% glucose. This solution was transferred to each well of a 96-well microtiter plate and incubated at 37°C for 24 h to permit biofilm formation on the well surface. Bacteria were removed and wells were washed twice with PBS. Compounds 1, 2, or vancomycin were added (in triplicate) to wells and serially diluted. The microtiter plate was then incubated at 37°C for 24 h. The plate was washed twice by submerging in deionized water. 0.1% (w/v) crystal violet was added to each well and allowed to stain the biofilm for 20 min before addition of 95% ethanol to decolorize. Using a kinetic microplate reader (Molecular Devices, Sunnyvale, CA, USA), the optical density of each well at 595 nm was measured. Percent biofilm mass reduction was calculated for each treatment regimen as compared to the control (wells receiving no treatment).