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Yeast Surface Display for Antibody Isolation: Library Construction, Library Screening, and Affinity Maturation

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Monoclonal Antibodies

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1131))

Abstract

Antibodies play key roles as reagents, diagnostics, and therapeutics in numerous biological and biomedical research settings. Although many antibodies are commercially available, oftentimes, specific applications require the development of antibodies with customized properties. Yeast surface display is a robust, versatile, and quantitative method for generating these antibodies and is accessible to single-investigator laboratories. This protocol details the key aspects of yeast surface display library construction and screening.

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References

  1. Gai SA, Wittrup KD (2007) Yeast surface display for protein engineering and characterization. Curr Opin Struct Biol 17:467–473

    Article  CAS  PubMed  Google Scholar 

  2. van Bloois E, Winter RT, Kolmar H et al (2011) Decorating microbes: surface display of proteins on Escherichia coli. Trends Biotechnol 29:79–86

    Article  PubMed  Google Scholar 

  3. Hoogenboom HR (2005) Selecting and screening recombinant antibody libraries. Nat Biotechnol 23:1105–1116

    Article  CAS  PubMed  Google Scholar 

  4. Winter G, Griffiths AD, Hawkins RE et al (1994) Making antibodies by phage display technology. Annu Rev Immunol 12:433–455

    Article  CAS  PubMed  Google Scholar 

  5. Lipovsek D, Plückthun A (2004) In-vitro protein evolution by ribosome display and mRNA display. J Immunol Methods 290:51–67

    Article  CAS  PubMed  Google Scholar 

  6. Boder ET, Wittrup KD (1997) Yeast surface display for screening combinatorial polypeptide libraries. Nat Biotechnol 15:553–557

    Article  CAS  PubMed  Google Scholar 

  7. Boder ET, Raeeszadeh-Sarmazdeh M, Price JV (2012) Engineering antibodies by yeast display. Arch Biochem Biophys 526:99–106

    Article  CAS  PubMed  Google Scholar 

  8. Chao G, Lau WL, Hackel BJ et al (2006) Isolating and engineering human antibodies using yeast surface display. Nat Protoc 1:755–768

    Article  CAS  PubMed  Google Scholar 

  9. Colby DW, Kellogg BA, Graff CP et al (2004) Engineering antibody affinity by yeast surface display. Methods Enzymol 388:348–358

    Article  CAS  PubMed  Google Scholar 

  10. Zaccolo M, Williams DM, Brown DM et al (1996) An approach to random mutagenesis of DNA using mixtures of triphosphate derivatives of nucleoside analogues. J Mol Biol 255:589–603

    Article  CAS  PubMed  Google Scholar 

  11. Rajan S, Sidhu SS (2012) Simplified synthetic antibody libraries. Methods Enzymol 502:3–23

    Article  CAS  PubMed  Google Scholar 

  12. de Wildt RMT, Mundy CR, Gorick BD et al (2000) Antibody arrays for high-throughput screening of antibody-antigen interactions. Nat Biotechnol 18:989–994

    Article  PubMed  Google Scholar 

  13. Pini A, Viti F, Santucci A et al (1998) Design and use of a phage display library. J Biol Chem 273:21769–21776

    Article  CAS  PubMed  Google Scholar 

  14. Feldhaus MJ, Siegel RW, Opresko LK et al (2003) Flow-cytometric isolation of human antibodies from a nonimmune Saccharomyces cerevisiae surface display library. Nat Biotechnol 21:163–170

    Article  CAS  PubMed  Google Scholar 

  15. Hermanson GT (2008) Bioconjugate techniques. Elsevier, Amsterdam

    Google Scholar 

  16. Beckett D, Kovaleva E, Schatz PJ (1999) A minimal peptide substrate in biotin holoenzyme synthetase-catalyzed biotinylation. Protein Sci 8:921–929

    Article  CAS  PubMed  Google Scholar 

  17. Wittrup KD (2002) Directed evolution of binding proteins by cell surface display: analysis of the screening process. In: Brakmann S, Johnsson K (eds) Directed molecular evolution of proteins. Wiley, New York, pp 111–126

    Chapter  Google Scholar 

  18. VanAntwerp JJ, Wittrup KD (2000) Fine affinity discrimination by yeast surface display and flow cytometry. Biotechnol Prog 16: 31–37

    Article  CAS  PubMed  Google Scholar 

  19. Hackel BJ, Kapila A, Wittrup KD (2008) Picomolar affinity fibronectin domains engineered utilizing loop length diversity, recursive mutagenesis, and loop shuffling. J Mol Biol 381:1238–1252

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  20. Swers JS, Kellogg BA, Wittrup KD (2004) Shuffled antibody libraries created by in vivo homologous recombination and yeast surface display. Nucleic Acids Res 32:e36

    Article  PubMed Central  PubMed  Google Scholar 

  21. Stemmer WPC (1994) Rapid evolution of a protein in vitro by DNA shuffling. Nature 370:389–391

    Article  CAS  PubMed  Google Scholar 

  22. Boder ET, Wittrup KD (1998) Optimal screening of surface-displayed polypeptide libraries. Biotechnol Prog 14:55–62

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

We thank Alessandro Angelini and Tiffany Chen for providing comments on the manuscript. This work was supported by CA96504, CA101830, and postdoctoral fellowship support to J.A.V. from the National Institutes of Health (Ruth L. Kirschstein National Research Service Award #1 F32 CA168057-01) and the Ludwig Fund for Cancer Research.

Author information

Authors and Affiliations

  1. Department of Chemical Engineering, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA

    James A. Van Deventer

  2. Department of Chemical and Biological Engineering, Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA, USA

    Karl Dane Wittrup

Authors
  1. James A. Van Deventer
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  2. Karl Dane Wittrup
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Editor information

Editors and Affiliations

  1. Département de Biochimie, University of Geneva, Geneva, Switzerland

    Vincent Ossipow

  2. NovImmune SA, Plan-les-Ouates, Switzerland

    Nicolas Fischer

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© 2014 Springer Science+Business Media, New York

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Van Deventer, J.A., Wittrup, K.D. (2014). Yeast Surface Display for Antibody Isolation: Library Construction, Library Screening, and Affinity Maturation. In: Ossipow, V., Fischer, N. (eds) Monoclonal Antibodies. Methods in Molecular Biology, vol 1131. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-992-5_10

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  • DOI: https://doi.org/10.1007/978-1-62703-992-5_10

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-991-8

  • Online ISBN: 978-1-62703-992-5

  • eBook Packages: Springer Protocols

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