title: High-throughgput phage-display screening in array format.
abstract: Emerging technologies for the design and generation of human antibodies require improved approaches enabling their screening, characterization and validation. Currently, strategies based on ELISA or western blot are used to that aim. However, the ever increasing number of novel antibodies generated would benefit from the development of new high-throughput (HT) platforms facilitating rapid antibody identification and characterization. Herein, we describe a protein chip bearing recombinant phage particles and based on a large phage antibody library. In this paper we have set forth a novel implementation which provides a powerful and simple methodology enabling the identification of single-chain variable fragments (scFv). As a proof-of-principle of this method, we tested it with recombinant antigen (human recombinant interleukin 8). Additionally, we developed a novel bioinformatics tool that serves to compare this novel strategy with traditional methods. The method described here, together with associated informatics tools, is robust, relatively fast and represents a step-forward in protocols including phage library screenings.
authors: Cancer Research Center, University of Salamanca-CSIC, IBSAL, Department of Medicine, General Service of Cytometry, Campus Miguel de Unamuno S/N, 37007 Salamanca, Spain; Proteomics Unit, Cancer Research Center, IBSAL, University of Salamanca-CSIC, Campus Miguel de Unamuno s/n, 37007 Salamanca, Spain.
authors: Paula
authors: P
authors: Díez
authors: ImmunoStep SL, Cancer Research Center, Avda, Universidad de Coimbra S/N, 37007 Salamanca, Spain.
authors: Ricardo
authors: R
authors: Jara-Acevedo
authors: Cancer Research Center, University of Salamanca-CSIC, IBSAL, Department of Medicine, General Service of Cytometry, Campus Miguel de Unamuno S/N, 37007 Salamanca, Spain; Proteomics Unit, Cancer Research Center, IBSAL, University of Salamanca-CSIC, Campus Miguel de Unamuno s/n, 37007 Salamanca, Spain.
authors: María
authors: M
authors: González-González
authors: Spanish National Research Council (CSIC)-Spanish National Biotechnology Centre (CNB), Darwin 3, Cantoblanco, 28049 Madrid, Spain.
authors: Juan
authors: J
authors: Casado-Vela
authors: Cancer Research Center, University of Salamanca-CSIC, IBSAL, Department of Medicine, General Service of Cytometry, Campus Miguel de Unamuno S/N, 37007 Salamanca, Spain; Proteomics Unit, Cancer Research Center, IBSAL, University of Salamanca-CSIC, Campus Miguel de Unamuno s/n, 37007 Salamanca, Spain.
authors: Noelia
authors: N
authors: Dasilva
authors: Cancer Research Center, University of Salamanca-CSIC, IBSAL, Department of Medicine, General Service of Cytometry, Campus Miguel de Unamuno S/N, 37007 Salamanca, Spain.
authors: Quentin
authors: Q
authors: Lécrevisse
authors: Cancer Research Center, University of Salamanca-CSIC, IBSAL, Department of Medicine, General Service of Cytometry, Campus Miguel de Unamuno S/N, 37007 Salamanca, Spain.
authors: Raquel
authors: R
authors: Bartolomé
authors: ImmunoStep SL, Cancer Research Center, Avda, Universidad de Coimbra S/N, 37007 Salamanca, Spain.
authors: José Carlos
authors: JC
authors: Claros
authors: ImmunoStep SL, Cancer Research Center, Avda, Universidad de Coimbra S/N, 37007 Salamanca, Spain.
authors: Alfredo
authors: A
authors: González
authors: ImmunoStep SL, Cancer Research Center, Avda, Universidad de Coimbra S/N, 37007 Salamanca, Spain.
authors: Ricardo
authors: R
authors: López
authors: Cancer Research Center, University of Salamanca-CSIC, IBSAL, Department of Medicine, General Service of Cytometry, Campus Miguel de Unamuno S/N, 37007 Salamanca, Spain.
authors: Alberto
authors: A
authors: Orfao
authors: Cancer Research Center, University of Salamanca-CSIC, IBSAL, Department of Medicine, General Service of Cytometry, Campus Miguel de Unamuno S/N, 37007 Salamanca, Spain; Proteomics Unit, Cancer Research Center, IBSAL, University of Salamanca-CSIC, Campus Miguel de Unamuno s/n, 37007 Salamanca, Spain. Electronic address: mfuentes@usal.es.
authors: Manuel
authors: M
authors: Fuentes
mesh_terms: D001483:Base Sequence; D061505:Cell Surface Display Techniques / Q000379:methods*; D003001:Cloning, Molecular; D019295:Computational Biology; D004247:DNA / Q000235:genetics; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D055767:Immobilized Proteins / Q000235:genetics / Q000276:immunology; D016209:Interleukin-8 / Q000235:genetics / Q000276:immunology; D008969:Molecular Sequence Data; D019151:Peptide Library; D040081:Protein Array Analysis / Q000379:methods; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D057127:Single-Chain Antibodies / Q000737:chemistry / Q000235:genetics
chemical_list: C508606:CXCL8 protein, human; D055767:Immobilized Proteins; D016209:Interleukin-8; D019151:Peptide Library; D011994:Recombinant Proteins; D057127:Single-Chain Antibodies; D004247:DNA
keywords: Antibody; Chip; High-throughput screening; Phage library; Phage-display array