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The input to the system can either be a contiguous protein sequence or a list of peptides, one per line.

The default selection on the webpage is "Protein sequence" (Figure 1 A), which means the input sequence is treated as a contiguous protein sequence (carriage returns and line breaks will be ignored). The PREDBALB/c input processing program decomposes the protein sequence into a series of 9-mer peptides overlapping by eight amino acids. Individual 9-mer peptides are then submitted for prediction. Predicted binding scores for all 9-mers are displayed in the result tables (Figure 1B). The 9-mer binding scores range form zero to ten, the higher the score the higher the probability of peptide being binder. PREDBALB/c has the option for plotting the binding scores of all the overlapping 9-mer peptides as a graph, in which X axis represents the start position of a 9-mer peptide and Y axis represents the binding score of the 9-mer peptide (Figure 1C). The user can sort the peptides by their binding scores and set a threshold to get binding regions composing by binding 9-mer peptides with binding scores above or equal to the user-set threshold.

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Figure 1. An example of the input page of PREDBALB/c when input is a single protein. The input protein sequence is Staphylococcal nuclease (Nase), the sequence type chosen is "Protein sequence" and the H2d alleles of interest are all H2d alleles. A) The input page. B) The main result page. The input sequence is decomposed into overlapping 9-mers for prediction of binding scores to each allele. C) Graphical view of the predicted binding scores to I-Ed molecule.

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When the input sequence type is set to "a list of peptide sequences", the input sequences separated by carriage returns or line breaks are treated as different peptides (Figure 2A). All overlapping 9-mers in each peptide are submitted for prediction. In the result tables, predicted binding scores to each allele are represented by the highest individual binding score of each input peptide to the allele. The predicted binding scores of individual 9-mers in each peptide in the list are not shown (Figure 2 B). To display the top scoring 9-mer peptides from each input peptide, the user can use the function "View binding peptides at threshold" (Figure 2 B). An explaination on choosing of threshold is displayed by clicking on the question mark on the webpage (http://research.i2r.a-star.edu.sg/predBalbc/HTML/specificity.html). In the result page (Figure 2 C), the 9-mers with binding scores equal to or above the thresholds at specificity level 0.90 are aligned with the input peptides. The predicted 9-mers are displayed with the names of the H2d alleles to which the 9-mer binding scores are above the threshold. According to the table in http://research.i2r.a-star.edu.sg/predBalbc/HTML/threshold.html, at specificity level 0.90, the threshold is 4.32 for Dd and 3.43 for Ld. In the first input peptide, YPILPEYLQCVK, there are 2 9-mer binders to Ld, YPILPEYLQ and LPEYLQCVK, and one binder to Dd, ILPEYLQCV.

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Figure 2. An example of the input page of PREDBALB/c when input is a list of pepitdes. The input is a list of peptides from chicken ovalbumin and the H2d alleles of interest are H2 class II alleles. A) The input page. B) The main result page. All 9-mers in each peptide are submitted for prediction. The predicted binding scores to each allele are represented by the highest individual binding score of each input peptide to the allele C) The alignment view of the predicted 9-mer binding peptides at threshold 8.

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