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Biopanning and rapid analysis of selective interactive ligands

Nature Medicine volume 7, pages 1249–1253 (2001)Cite this article

Abstract

Here we introduce a new approach for the screening, selection and sorting of cell-surface–binding peptides from phage libraries. Biopanning and rapid analysis of selective interactive ligands (termed BRASIL) is based on differential centrifugation in which a cell suspension incubated with phage in an aqueous upper phase is centrifuged through a non-miscible organic lower phase. This single-step organic phase separation is faster, more sensitive and more specific than current methods that rely on washing steps or limiting dilution. As a proof-of-principle, we screened human endothelial cells stimulated with vascular endothelial growth factor (VEGF) and constructed a peptide-based ligand-receptor map of the VEGF family. Next, we validated the motif PQPRPL as a novel chimeric ligand mimic that binds specifically to VEGF receptor-1 and to neuropilin-1. BRASIL may prove itself a superior method for probing target cell surfaces with a broad range of potential applications.

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Figure 1: Cell-binding assays with a defined αv integrin phage ligand and phage-cell–binding optimization with BRASIL.
Figure 2: Screening VEGF165-stimulated HUVECs by BRASIL.
Figure 3: Binding of the CPQPRPLC peptide to immobilized VEGF receptors.

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Acknowledgements

This work was supported by the NIH (CA90270 and CA8297601 to RP; CA90270 and CA9081001 to W.A.) and by a Gillson-Longenbaugh Foundation Award (to R.P. and W.A.). R.J.G. was supported by FAPESP (Brazil), M.C.V. by the Department of Defense and J.L. by the Susan G. Komen Breast Cancer Foundation.

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  1. The University of Texas M.D. Anderson Cancer Center, Houston, Texas, USA

    Ricardo J. Giordano, Marina Cardó-Vila, Johanna Lahdenranta, Renata Pasqualini & Wadih Arap

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  1. Ricardo J. Giordano
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  2. Marina Cardó-Vila
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  3. Johanna Lahdenranta
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  4. Renata Pasqualini
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  5. Wadih Arap
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Correspondence to Renata Pasqualini or Wadih Arap.

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Giordano, R., Cardó-Vila, M., Lahdenranta, J. et al. Biopanning and rapid analysis of selective interactive ligands. Nat Med 7, 1249–1253 (2001). https://doi.org/10.1038/nm1101-1249

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