Mts compound

Cells were seeded in 96-well plate in triplicate at 3 × 10³ per well and cultured in the presence or absence of cisplatin or the IKKβ inhibitor at the indicated concentrations and time course. At the end of each time point, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) compound (Promega) was added for 1 hours at 37°C. Colorimetric readouts were read at 490 nm on a Versamax Microplate Reader (Molecular Devices).

Cells were seeded at 2000 or 3000 cells per well in 96-well plates, then treated with DMSO or Compound A daily as indicated. At each time point, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxy-phenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) compound (Promega) was added and absorbance was read at 490 nm on a Versamax Microplate Reader (Molecular Devices).

Cells were plated at 1,500–2,500 cells/well in 96-well plates in growth media with FBS for proper cell adherence. In each experiment, cells were monitored by phase contrast microscopy to assess attachment and overall health. Assays in separate plates were stopped at 0–5 days after plating. The quantity of viable cells was assessed with a 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) compound (Promega, Madison, WI, USA) after 1 h incubation at 37°C. MTS is bioreduced by metabolically active cells into a colored formazan product soluble in tissue culture media, which can be read at 490 nm absorbance. The absorbance is directly proportional to the number of living cells. In every experiment, a standard curve was constructed for each line relating absorbance to cell number. Results were analyzed in terms of cell number.
In some experiments, cells were treated at the start of the growth period with 1–200 µM 8-(4-chlorophenylthio)-adenosine-3′,5′-cyclic monophosphate (8-CPT-cAMP), 8-CPT-2′-O-methyl-cAMP (me-cAMP), or N-6-phenyl-cAMP (phe-cAMP). 8-CPT-cAMP is a non-selective agonist, while me-cAMP and phe-cAMP are selective for EPAC and protein kinase A (PKA), respectively.