Metabolism activity assessment using the resazurin test

The resazurin test is a colorimetric assay for cell viability that involves the reduction of resazurin to resorufin by the metabolically active mitochondria of cells. Resorufin is a fluorescent compound that exhibits a pink colouring. After a time of incubation, the amount of resorufin formed is directly proportional to the number of metabolically active cells. Cells cultured in 2D or 3D treated with the drugs for 5 days at 37°C were washed with PBS^* and incubated with 60 μM of resazurin solubilized in PBS. Optical densities were measured at 570 and 620 nm after 6 to 15 hours of incubation. The amount of reduced compound (resorufin) corresponding to the number of active cells per well was calculated for each condition. The survival rate was determined considering the ratio (OD_570-620nm treated well/OD_570-620nm untreated control) x 100. The half maximal inhibitory concentration (I_C50) was calculated with the following formula: I_C50 = EXP (LN (concentration > 50% inhibition) - ((signal > 50% inhibition - 50) / (signal > 50% inhibition - signal < 50% inhibition) ^* LN (concentration >50% inhibition / concentration <50% inhibition))).