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

Author(s): Zhi Duan, Henrik Siegumfeldt.

Journal Name: Combinatorial Chemistry & High Throughput Screening

Volume 13 , Issue 9 , 2010

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Abstract:

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 αs1-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.

Keywords: Phage display, scFv, Tomlinson I + J libraries, casein, peptide.

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Article Details

VOLUME: 13
ISSUE: 9
Year: 2010
Page: [818 - 828]
Pages: 11
DOI: 10.2174/138620710792927376
Price: $58

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