Figure 2
From: Mapping polyclonal antibody responses to bacterial infection using next generation phage display
Screening of peptides specific for S. Enteritidis infection in chickens.
Peptides identified in silico as being enriched against IgY from chickens experimentally infected with S. Enteritidis compared to birds infected with S. Hadar were assessed for binding to IgY from 19 birds infected with S. Enteritidis, 9 infected with S. Hadar and 16 infected with S. Typhimurium. Synthetic peptide was immobilised on maxisorb plates and probed with the IgY, bound antibody was then detected with an anti-IgY-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). ROC analysis for each peptide revealed 9 peptides (out of 15) with associated p values < 0.05 indicating that they are highly discriminatory for sera from S. Enteritidis infected chickens. Binding of IgY samples to these 9 peptides is shown (A) as being positive (grey boxes) or negative (white boxes). Animal numbers are shown and those in italics provided IgY for the phage panning. The most discriminatory individual peptides was AEGEFEPQQSARPS that bound to 7 of the 19 samples from S. Enteritidis infected chickens. ROC analysis is shown (B) along with the ELISA signals (C). This peptide had 100% specificity and 37% sensitivity, the positive and negative predicted values are shown for the samples tested.
