MHCI-peptide length preference is not determined by MHCI binding. T2 cells (0.5 × 10⁶) were incubated in RPMI 1640 medium with either 100μM HPVGEADYFEY (non-HLA A*0201 binder), 100μM GILGFVFTL (HLA A*0201 binder) or 1mM of the indicated sizing scan mixture (X⁸, X⁹, X¹⁰, X¹¹, X¹², or X¹³) at 26°C for 14-16 hours, then at 37°C for 2 hours, before staining for HLA A*0201 surface expression. The conditions used here are representative of those that were used for the experiments in Figure 1 and supplemental Figure 1. Duplicate samples were acquired for each condition using a FACSCanto II flow cytometer. Data were analyzed with FlowJo software. NP indicates no peptide. Error bars represent SDs.