title: Using phage and yeast display to select hundreds of monoclonal antibodies: application to antigen 85, a tuberculosis biomarker.
abstract: BACKGROUND
  Current diagnostic methods for tuberculosis (TB), a major global health challenge that kills nearly two million people annually, are time-consuming and inadequate. During infection a number of bacterial molecules that play a role in the infective process are released and have been proposed as biomarkers for early TB diagnosis. Antigen 85 (Ag85) is the most abundant secreted TB protein, and a potential target for this diagnostic approach. One of the bottlenecks in the direct detection of such bacterial targets is the availability of robust, sensitive, specific antibodies.
  
  
  METHODS
  Using Ag85 as a model, we describe a method to select antibodies against any potential target using a novel combination of phage and yeast display that exploits the advantage of each approach.
  
  
  RESULTS
  The efficiency of this approach was attested to by the 111 specific antibodies identified in initial screens. These were assessed for binding to the different Ag85 subunits, affinity, and activity in sandwich assays.
  
  
  CONCLUSIONS
  The novelty of this approach lies in the possibility of screening the entire output of a phage antibody selection in a single experiment by yeast display. This can be considered analogous to carrying out a million ELISAs. The monoclonal antibodies (mAbs) identified in this way show high binding affinity and selectivity for the antigens and offer an advantage over traditional mAbs produced by relatively expensive and time consuming techniques. This approach has wide applicability, and the affinity of selected antibodies can be significantly improved, if required.
authors.affiliation: Bioscience Division, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America.
authors.forename: Fortunato
authors.initials: F
authors.lastname: Ferrara
authors.forename: Leslie A
authors.initials: LA
authors.lastname: Naranjo
authors.forename: Sandeep
authors.initials: S
authors.lastname: Kumar
authors.forename: Tiziano
authors.initials: T
authors.lastname: Gaiotto
authors.forename: Harshini
authors.initials: H
authors.lastname: Mukundan
authors.forename: Basil
authors.initials: B
authors.lastname: Swanson
authors.forename: Andrew R M
authors.initials: AR
authors.lastname: Bradbury
mesh_terms: D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology*; D000915:Antibody Affinity / Q000276:immunology; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology*; D015415:Biomarkers; D061505:Cell Surface Display Techniques; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000276:immunology*; D019151:Peptide Library; D011485:Protein Binding / Q000276:immunology; D055550:Protein Stability; D021122:Protein Subunits / Q000737:chemistry / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000276:immunology; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology; D014376:Tuberculosis / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D015415:Biomarkers; D019151:Peptide Library; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies