Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Isolation of engineered, full-length antibodies from libraries expressed in Escherichia coli


We describe facile isolation of full-length IgG antibodies from combinatorial libraries expressed in E. coli. Full-length heavy and light chains are secreted into the periplasm, where they assemble into aglycosylated IgGs that are captured by an Fc-binding protein that is tethered to the inner membrane. After permeabilizing the outer membrane, spheroplast clones expressing so-called E-clonal antibodies, which specifically recognize fluorescently labeled antigen, are selected using flow cytometry. Screening of a library constructed from an immunized animal yielded several antibodies with nanomolar affinities toward the protective antigen of Bacillus anthracis.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Prices vary by article type



Prices may be subject to local taxes which are calculated during checkout

Figure 1: The E-clonal technology.
Figure 2: Analysis of IgG clones selected from the anti-PA library.


  1. Kohler, G. & Milstein, C. Nature 256, 495–497 (1975).

    Article  CAS  Google Scholar 

  2. Li, J. et al. Proc. Natl. Acad. Sci. USA 103, 3557–3562 (2006).

    Article  CAS  Google Scholar 

  3. Fishwild, D.M. et al. Nat. Biotechnol. 14, 845–851 (1996).

    Article  CAS  Google Scholar 

  4. Winter, G., Griffiths, A.D., Hawkins, R.E. & Hoogenboom, H.R. Annu. Rev. Immunol. 12, 433–455 (1994).

    Article  CAS  Google Scholar 

  5. Boder, E.T. & Wittrup, K.D. Nat. Biotechnol. 15, 553–557 (1997).

    Article  CAS  Google Scholar 

  6. Daugherty, P.S., Olsen, M.J., Iverson, B.L. & Georgiou, G. Protein Eng. 12, 613–621 (1999).

    Article  CAS  Google Scholar 

  7. Lipovsek, D. & Pluckthun, A. J. Immunol. Methods 290, 51–67 (2004).

    Article  CAS  Google Scholar 

  8. Harvey, B.R. et al. Proc. Natl. Acad. Sci. USA 101, 9193–9198 (2004).

    Article  CAS  Google Scholar 

  9. Hoogenboom, H.R. Nat. Biotechnol. 23, 1105–1116 (2005).

    Article  CAS  Google Scholar 

  10. Jendeberg, L. et al. J. Mol. Recognit. 8, 270–278 (1995).

    Article  CAS  Google Scholar 

  11. Simmons, L.C. et al. J. Immunol. Methods 263, 133–147 (2002).

    Article  CAS  Google Scholar 

  12. Chen, G., Dubrawsky, I., Mendez, P., Georgiou, G. & Iverson, B.L. Protein Eng. 12, 349–356 (1999).

    Article  CAS  Google Scholar 

  13. Nilsson, B. et al. Protein Eng. 1, 107–113 (1987).

    Article  CAS  Google Scholar 

  14. Maynard, J.A. et al. Nat. Biotechnol. 20, 597–601 (2002).

    Article  CAS  Google Scholar 

  15. Tao, M.H. & Morrison, S.L. J. Immunol. 143, 2595–2601 (1989).

    CAS  PubMed  Google Scholar 

Download references


We thank Clinton E. Leysath for preparation of the cDNA from splenocytes of PA-immunized mice. This work was supported by the Foundation for Research.

Author information

Authors and Affiliations


Corresponding author

Correspondence to George Georgiou.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Expression and purification of intact IgGs in the bacterial periplasm. (PDF 463 kb)

Supplementary Fig. 2

Binding of IgG to spheroplasts displaying the protein ZZ domain. (PDF 63 kb)

Supplementary Fig. 3

Evaluation of the NlpA-ZZ-IgG complex stability. (PDF 48 kb)

Supplementary Fig. 4

Construction of immunized anti-PA IgG Library. (PDF 30 kb)

Supplementary Fig. 5

Amino acid sequence analysis of the isolated anti-PA IgG individual clones. (PDF 15 kb)

Supplementary Fig. 6

An example of kinetic analysis of YMF10 IgG toward PA-63 using an IgG capture method. (PDF 46 kb)

Supplementary Methods (PDF 48 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Mazor, Y., Van Blarcom, T., Mabry, R. et al. Isolation of engineered, full-length antibodies from libraries expressed in Escherichia coli. Nat Biotechnol 25, 563–565 (2007).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing