TGEM was performed as outlined in the Methods testing seven allele
types. PBMCs were cultured in the presence of pools of five MCPyV 20-mer
peptides (WED, GNI, NSG, PNG; amino acid sequence indicated) for 10–14
days, followed by staining for pools of tetrameric complexes of 10 peptides
(composed of two combined pools) bound to HLA class II molecules. After
identification of CD4+tetramer+ T cells, the same cultures
were stained with single tetramers containing each relevant peptide within the
positive pool. Representative flow plots are shown for each peptide. For each
epitope, four individuals were tested, and one to four persons were positive as
detailed in column 7 of Table 1. The
percentage of viable CD4+tetramer+ T cells (negative for
CD8/CD14/CD19) in the lymphocyte forward/side scatter region are denoted.