Supplementary Figure 2: Immunopeptidomic analysis of human and viral peptides.

(a) Infection and viability rates for the immunopeptidomics experiments. (b) Experimental outline for immunopeptidomics analysis of IBV-infected C1R cells. Lysates were generated from C1R parental or C1R.A*02:01 cells and the transfected HLA-A*02:01, endogenous (parental) class I and class II molecules immunoaffinity purified sequentially using BB7.2 (HLA-A2 specific), w6/32 (pan class I) and a mixture of LB3.1 (HLA-DR), SPV-L3 (HLA-DQ) and B721 (HLA-DP). Peptides were eluted from the purified HLA and interrogated by LC–MS/MS using an information dependent acquisition (IDA) strategy and preliminary lists of HLA ligands generated for HLA-A*02:01, parental class I and class II. HLA-A*02:01 ligands were further refined by filtering for sequences in C1R parental purifications from all antibodies (represent non-specific pull down by BB7.2, and contamination with endogenous HLA molecules) as well as sequences in C1R.A*02:01 LB3.1/SPV-L3/B721 purifications (represent contamination with endogenous class II molecules). Peptides from C1R.A*02:01 w6/32 purifications were not included in the contaminant list due to the potential for excess HLA-A*02:01 not bound to the preceding BB7.2 resin to be isolated by w6/32. (c) Number of peptides identified per condition and experiment. (d) Distribution of IBV-derived HLA ligands across the B/Malaysia proteome, distinguished as likely parental class I (HLA-B*35:03 and HLA-C*04:01) or HLA class II ligands. All immunopeptidomic data are from two independent experiments.