Profiling the autoantibody repertoire by screening phage-displayed human cDNA libraries

Methods Mol Biol. 2009:570:353-69. doi: 10.1007/978-1-60327-394-7_20.

Abstract

The advent of the serological identification of antigens by procedures such as cDNA cloning and recombinant protein expression has allowed the direct molecular definition of immunogenic proteins. The phage-display technology provides several advantages over conventional immunoscreening procedures based on plasmid or lambda-phage cDNA libraries. So far, attempts to display open reading frames, such as those encoded by cDNA fragments, on filamentous phages have not been very successful. We managed to develop a strategy based on "folding reporters" which allows filtering out open reading frames from DNA and displaying them on filamentous phages in such a way that they are amenable to subsequent selection or screening. Once the cDNA library of interest is created, phage-display technology is used for selection of novel putative antigens; these are then validated by printing isolated protein on microarray and screening with patients' sera.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Review

MeSH terms

  • Autoantibodies / analysis*
  • Autoantibodies / genetics
  • Autoantibodies / metabolism
  • Gene Library*
  • High-Throughput Screening Assays / methods
  • Humans
  • Models, Biological
  • Open Reading Frames
  • Peptide Library*
  • Protein Array Analysis / methods
  • Recombinant Proteins / analysis
  • Recombinant Proteins / immunology

Substances

  • Autoantibodies
  • Peptide Library
  • Recombinant Proteins