The percentage inhibition was plotted against the drug concentration by calculating the difference between the geometric means of test wells and negative wells (without drug). The result was divided by the difference in the geometric mean between the positive control wells (at maximum drug concentration) and negative wells (without drug). Inhibition curves were defined using a sigmoid dose–response curve with a variable slope (GraphPad Prism v5.0 software). The fold change (FC) in drug susceptibility was determined by dividing the IC50 for every protease-recombinant plasmid sample by the IC50 for the tipranavir- or darunavir-sensitive protease-recombinant plasmid (pcDNA3_GFP_PR-wt).