Figure 1
From: Mapping polyclonal antibody responses to bacterial infection using next generation phage display
Screening of peptides specific for S. Typhimurium infection in pigs.
Peptides identified in silico as being enriched against IgG from pigs experimentally infected with S. Typhimurium were assessed for binding to sera antibodies from 24 animals taken before challenge (pre-infection) and after challenge (post-infection). Synthetic peptide was immobilised on maxisorb plates and probed with the sera, bound antibody was then detected with an anti-porcine-IgG-AP conjugate. All samples were analysed in duplicate, ELISA signals after 2 hours incubation with substrate were taken. Binding was defined by applying a cut-off value for each peptide that was determined as the mean +3SD of the signals for the 24 non-infected samples (dashed line in C,E). ROC analysis for each peptide revealed 22 peptides (out of 27) with associated p values < 0.05 indicating that they are highly discriminatory for sera from infected pigs. Binding of sera samples to these 22 peptides is shown (A) as being positive (grey boxes) or negative (white boxes). Animal numbers are shown and those in italics provided sera samples for the phage panning. Analysis of the 22 peptides showed binding to all 24 sera samples from infected pigs but not to any sera from non-infected pigs. Examples of binding of individual peptides are also shown for AEGEFVQATDTNS (B,C) and AEGEFPLHNGNERL (D,E) and both bound to 13 of the 24 samples from infected pigs. ROC analysis is shown (B,D) along with the ELISA signals for each of the 24 infected and non-infected animals binding to peptides (C,E). These peptides had 100% specificity and 54% sensitivity, the positive and negative predicted values are shown for the samples tested.
