Cell Viability and Analysis for Single and Combination Drug Screening

Cell viability was analyzed on Day 7 using the MTT assay as described earlier. Day 7 response data and synergy for single and combination drug studies were evaluated using the Chou-Talalay method of synergy testing via CalcuSyn⁴⁸. In Microsoft Excel, the raw MTT absorbance data from quadruplicate samples were normalized to the DMSO control. Then, to determine effect, normalized absorbances were subtracted from 1. The quadruplicate sample values were then averaged together to obtain a mean. These effect data were entered into CalcuSyn (v. 2.1) as single doses and combinations (non-constant ratios). Combination Indices (CI) were generated. CI values and their indications are as follows: <1 is synergistic, =1 is additive, and >1 is antagonistic. In cases where neither single nor combination dosing resulted in an inhibitory effect less than 25% of control levels, CI scores were not determined and reported as “not a number” (NaN)⁴⁹.