Skip to main content

Antibody Epitope Mapping Using SPOT™ Peptide Arrays

  • Protocol
  • First Online:
Epitope Mapping Protocols

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 524))

Summary

Information at the amino acid level about the epitopes of proteins recognized by antibodies or antibody fragments is important for their use as biological and diagnostic tools, therapeutic molecules, and for understanding molecular recognition events in general. The use of chemically prepared arrays of short peptides has emerged as a powerful tool to identify and characterize antibody epitopes. In this chapter the SPOT™ synthesis technique is described in detail. In addition, three different types of peptide libraries and their applications are described: protein sequence-derived scans of overlapping peptides (peptide scans) used to locate epitopes within the protein sequence, truncation libraries used to identify the minimal peptide length required for antibody binding, and complete substitutional analyses to identify the key residues important for antibody binding.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 109.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Frank, R. (1992) Spot synthesis: an easy technique for the positionally addressable, parallel chemical synthesis on a membrane support. Tetrahedron 48, 9217–9232.

    Article  CAS  Google Scholar 

  2. Frank, R. (Date of filing: 31.08.1990) Verfahren zur parallelen Herstellung von trgergebundenen oder freien Peptiden, trgergebundene Peptide and ihre Verwendung. DE 4027675

    Google Scholar 

  3. Geysen, H. M., Rodda, S. J., Mason, T. J., Tribbick, G., and Schofs, P. G. (1987) Strategies for epitope analysis using peptide synthesis. J. Immunol. Methods 102, 259–274.

    Article  PubMed  CAS  Google Scholar 

  4. Geysen, H. M. (Date of filing: 08. 03. 1984) Method for determining antigenically active amino acids sequences. EP 0138855.

    Google Scholar 

  5. Pinilla, C., Appel, J. R., and Houghten, R. (1993) Functional importance of amino acid residues making up antigenic determinants. Mol. Immunol. 30, 577–585.

    Article  PubMed  CAS  Google Scholar 

  6. Molina, F., Laune, D., Gougat, C., Pau, B., and Granier, C. (1996) Improved performances of spot multiple peptide synthesis. Peptide Res. 9, 151–155.

    CAS  Google Scholar 

  7. Reineke, U., Kramer, A., and Schneider-Mergener, J. (1999) Antigen sequence- and library-based mapping of linear and discontinuous protein-protein-interaction sites by spot synthesis. Curr. Top. Microbiol. Immunol. 243, 23–36.

    Article  PubMed  CAS  Google Scholar 

  8. Wenschuh, H., Volkmer-Engert, R., Schmidt, M., Schulz, M., Schneider-Mergener J., and Reineke U. (2000) Coherent membrane supports for parallel microsynthesis and screening of bioactive peptides. Biopolymers (Peptide Science) 55, 188–206.

    Article  CAS  Google Scholar 

  9. Reineke, U., Volkmer-Engert R., and Schneider-Mergener, J. (2001) Applications of peptide arrays prepared by the SPOT-technology. Curr. Opin. Biotechnol. 12, 59–64.

    Article  PubMed  CAS  Google Scholar 

  10. Frank, R. and Schneider-Mergener, J. (2002) SPOT-synthesis: scope and applications, in Peptide Arrays on Membrane Supports – Synthesis and Applications, Springer Laboratory Manual (Koch, J., and Mahler, M., eds.), Springer-Verlag, Heidelberg, Germany, pp.1–22.

    Google Scholar 

  11. Reimer, U., Reineke, U., and Schneider-Mergener, J. (2002) Peptide arrays: from macro to micro. Curr. Opin. Biotechnol. 13, 315–320.

    Article  PubMed  CAS  Google Scholar 

  12. Reineke, U., Schneider-Mergener, J., and Schutkowski, M. (2006) Peptide arrays in proteomics and drug discovery, in BioMEMS and Biomedical Nanotechnology, Volume II Micro/Nano Technology for Genomics and Proteomics (Ozkan, M., and Heller, M. J., eds.), Springer-Verlag, Berlin, pp. 161–282.

    Google Scholar 

  13. Ast, T., Heine, N., Germeroth, L., Schneider-Mergener, J., and Wenschuh, H. (1999) Efficient assembly of peptomers on continuous surfaces. Tetrahedron Lett. 40, 4317–4318.

    Article  CAS  Google Scholar 

  14. Van Regenmortel, M. H. V. (1994) The recognition of proteins and peptides by antibodies, in Immunochemistry (van Oss, C. J., and van Regenmortel, M. H. V., eds.), Marcel Dekker, New York, NY, pp. 277–300.

    Google Scholar 

  15. Kramer, A., Reineke, U., Dong, L., Hoffmann, B., Hoffmüller, U., Winkler, D., Volkmer-Engert, R., and Schneider-Mergener, J. (1999) Spot synthesis: observations and optimizations. J. Peptide Res. 54, 319–327.

    Article  CAS  Google Scholar 

  16. Wilson, M. B. and Nakane, P. K. (1978) Recent developments in the periodate method of conjugating horseradish peroxidase (HRPO) to antibodies, in Immunofluorescence Related Staining Techniques (Knapp, W., Holubar, K., and Wick, G., eds.), Elsevier, Amsterdam, pp. 215–224.

    Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Ulrich Reineke .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2009 Humana Press, a part of Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Reineke, U., Sabat, R. (2009). Antibody Epitope Mapping Using SPOT™ Peptide Arrays. In: Schutkowski, M., Reineke, U. (eds) Epitope Mapping Protocols. Methods in Molecular Biology™, vol 524. Humana Press. https://doi.org/10.1007/978-1-59745-450-6_11

Download citation

  • DOI: https://doi.org/10.1007/978-1-59745-450-6_11

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-934115-17-6

  • Online ISBN: 978-1-59745-450-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics