Flowchart of down-selection of putative MHC binders for experimental evaluation.

<p>The whole-genome immunoinformatic analysis conducted on the 1754 <i>F. tularensis</i> ORF products and the downstream selection of the various subsets of peptides is provided. Boxes shaded in light blue: Subsets I and II were described in a previous study <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036440#pone.0036440-Zvi1" target="_blank">[14]</a> and are added for the sake of completeness. Boxes shaded in cyan: Subsets III–V of peptides selected and evaluated in this study. In all down selection steps referred to as : “Selected for evaluation" (in Subset III, IV and V), clusters or peptides are selected at random as an operational filter for reduction of number of peptides to be experimentally tested, unlike Subset II, were selection of peptides was at random with respect to clusters. Subset III contains 401 peptides selected from clusters having density of 0.8 up to 1.0 (1.0 not included); Subset IV is composed of 200 peptides selected from the overall 3016 predicted MHC binders (IC₅₀< = 500 nM) located outside the high-density cluster regions in the 41 source proteins of Subset I (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0036440#pone-0036440-g002" target="_blank">Figure 2</a>); Subset V includes 92 predicted epitopes having an IC₅₀ of 10 nM and lower, representing putative high-affinity MHC binders.