An Efficient Method for Isolating Antibody Fragments Against Small Peptides by Antibody Phage Display

Zhi Duan, Henrik Siegumfeldt

Research output: Contribution to journalJournal articleResearchpeer-review


We generated monoclonal scFv (single chain variable fragment) antibodies from an antibody phage display library towards three small synthetic peptides derived from the sequence of alpha(s1)-casein. Key difficulties for selection of scFv-phages against small peptides were addressed. Small peptides do not always bind efficiently to passive adsorption surfaces, and we developed a simple method to quantify the binding capacity of surfaces with the peptides. Background binding (the binding of scFvs to the background matrix) is an obstacle for successful selection, and we evaluated two methods that drastically reduced the background binding. An optimized method therefore enabled a panning procedure where the specific (peptide binding) scFv-phages were always dominant. Using 15-mer peptides immobilized on Nunc Immobilizer Streptavidin plates, we successfully generated scFvs specifically against them. The scFvs were sequenced and characterized, and specificity was characterized by ELISA. The methods developed in this study are universally applicable for antibody phage display to efficiently produce antibody fragments against small peptides.
Original languageEnglish
JournalCombinatorial Chemistry & High Throughput Screening
Issue number9
Pages (from-to)818-828
Publication statusPublished - 2010
Externally publishedYes


  • Facultatively Anaerobic Gram-Negative Rods Eubacteria Bacteria Microorganisms (Bacteria, Eubacteria, Microorganisms) - Enterobacteriaceae [06702] Escherichia coli species strain-TG1, strain-KM13
  • alpha-s1-casein
  • single chain variable fragment antibody scFv antibody
  • small peptide
  • streptavidin 9013-20-1
  • synthetic peptide
  • 10060, Biochemistry studies - General
  • 31000, Physiology and biochemistry of bacteria
  • enzyme linked immunosorbent assay ELISA laboratory techniques, immunologic techniques
  • phage antibody display laboratory techniques, immunologic techniques
  • Biochemistry and Molecular Biophysics
  • Methods and Techniques


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