Steady glo luciferase assay system reagent

VK2-HRE and RAW-HRE cells were plated the day before in white opaque, tissue culture 96 well plates in culture media as described above. Cells were pretreated with 12 μg/mL AKB-4924 or vehicle for 4 h before infection. Cells were infected with GBS COH1 or isogenic ΔcylE at MOI = 20 as indicated in figure legends. At 2 h postinfection, cells were washed once with PBS and lysed with 25 μL/well of Steady-Glo® Luciferase Assay System reagent (Promega) for 5 min at room temp. RPMI-1640, containing no phenol red, was added (25 μL/well), mixed, incubated at room temperature for 15 min, and luminescence was then measured on an EnSpire Alpha Multimode Plate Reader (PerkinElmer). Arbitrary units of luminescence were normalized across individual experiments using values from vehicle-treated, uninfected VK2-HRE or RAW-HRE cells.