TY - JOUR
T1 - Antibody biomarker discovery through in vitro directed evolution of consensus recognition epitopes
AU - Ballew, John T.
AU - Murray, Joseph A.
AU - Collin, Pekka
AU - Mäki, Markku
AU - Kagnoff, Martin F.
AU - Kaukinen, Katri
AU - Daugherty, Patrick S.
PY - 2013/11/26
Y1 - 2013/11/26
N2 - To enable discovery of serum antibodies indicative of disease and simultaneously develop reagents suitable for diagnosis, in vitro directed evolution was applied to identify consensus peptides recognized by patients' serum antibodies. Bacterial cell-displayed peptide libraries were quantitatively screened for binders to serum antibodies from patients with celiac disease (CD), using cellsorting instrumentation to identify two distinct consensus epitope families specific to CD patients (PEQ and E/DxFVY/FQ). Evolution of the E/DxFVY/FQ consensus epitope identified a celiac-specific epitope, distinct from the two CD hallmark antigens tissue transglutaminase- 2 and deamidated gliadin, exhibiting 71% sensitivity and 99% specificity (n = 231). Expansion of the first-generation PEQ consensus epitope via in vitro evolution yielded octapeptides QPEQAFPE and PFPEQxFP that identified ω- and γ-gliadins, and their deamidated forms, as immunodominant B-cell epitopes in wheat and related cereal proteins. The evolved octapeptides, but not first-generation peptides, discriminated one-way blinded CD and non-CD sera (n = 78) with exceptional accuracy, yielding 100% sensitivity and 98% specificity. Because this method, termed antibody diagnostics via evolution of peptides, does not require prior knowledge of pathobiology, it may be broadly useful for de novo discovery of antibody biomarkers and reagents for their detection.
AB - To enable discovery of serum antibodies indicative of disease and simultaneously develop reagents suitable for diagnosis, in vitro directed evolution was applied to identify consensus peptides recognized by patients' serum antibodies. Bacterial cell-displayed peptide libraries were quantitatively screened for binders to serum antibodies from patients with celiac disease (CD), using cellsorting instrumentation to identify two distinct consensus epitope families specific to CD patients (PEQ and E/DxFVY/FQ). Evolution of the E/DxFVY/FQ consensus epitope identified a celiac-specific epitope, distinct from the two CD hallmark antigens tissue transglutaminase- 2 and deamidated gliadin, exhibiting 71% sensitivity and 99% specificity (n = 231). Expansion of the first-generation PEQ consensus epitope via in vitro evolution yielded octapeptides QPEQAFPE and PFPEQxFP that identified ω- and γ-gliadins, and their deamidated forms, as immunodominant B-cell epitopes in wheat and related cereal proteins. The evolved octapeptides, but not first-generation peptides, discriminated one-way blinded CD and non-CD sera (n = 78) with exceptional accuracy, yielding 100% sensitivity and 98% specificity. Because this method, termed antibody diagnostics via evolution of peptides, does not require prior knowledge of pathobiology, it may be broadly useful for de novo discovery of antibody biomarkers and reagents for their detection.
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U2 - 10.1073/pnas.1314792110
DO - 10.1073/pnas.1314792110
M3 - Article
C2 - 24222690
AN - SCOPUS:84888385917
SN - 0027-8424
VL - 110
SP - 19330
EP - 19335
JO - Proceedings of the National Academy of Sciences of the United States of America
JF - Proceedings of the National Academy of Sciences of the United States of America
IS - 48
ER -