Screening technologies for recombinant antibody libraries

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Mieko Kato Yoshiro Hanyu


Monoclonal antibodies have become increasingly important in research and diagnosis, and therapeutics. Many antibody preparations are on the market, which has grown to an astounding size. In diagnosis, the use of antibodies has helped to enable early diagnoses of a wide variety of illnesses. The ability to establish antibodies to various types of antigens has accelerated proteomics research. The antibodies with higher affinity and specificity are required in each field, but these needs cannot be met with conventional monoclonal production technology, thus necessitating monoclonal antibody production using recombination technology. The use of recombinant technology enables the production of antibodies to the antigens for which antibodies cannot be produced using conventional production technology. In vitro selection techniques enable screening in the environment in which antibodies are used, thus in turn enabling the establishment of antibodies suited to specific uses. In the production of recombinant monoclonal antibodies, it is important to prepare a highly diverse library in order to identify positive clones using screening technology with little background reaction. The present paper explains screening technology in recombinant monoclonal antibody production, demonstrates the problems and issues in production, and discusses the future outlook of this technology.

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How to Cite
KATO, Mieko; HANYU, Yoshiro. Screening technologies for recombinant antibody libraries. Medical Research Archives, [S.l.], v. 2, n. 7, nov. 2015. ISSN 2375-1924. Available at: <>. Date accessed: 22 july 2018.
recombinant antibody; library; screening; single-chain variable fragment; phage display; panning
Review Articles


Accardi, L., & Di Bonito, P. (2010). Antibodies in single-chain format against tumour-associated antigens: present and future applications. Current Medicinal Chemistry, 17(17), 1730–55. Retrieved from

Akahori, Y., Kurosawa, G., Sumitomo, M., Morita, M., Muramatsu, C., Eguchi, K., … Kurosawa, Y. (2009). Isolation of antigen/antibody complexes through organic solvent (ICOS) method. Biochemical and Biophysical Research Communications, 378(4), 832–5. doi:10.1016/j.bbrc.2008.11.129

Buhr, D. L., Acca, F. E., Holland, E. G., Johnson, K., Maksymiuk, G. M., Vaill, A., … Kiss, M. M. (2012). Use of micro-emulsion technology for the directed evolution of antibodies. Methods (San Diego, Calif.), 58(1), 28–33. doi:10.1016/j.ymeth.2012.07.007

Chan, C. E. Z., Chan, A. H. Y., Lim, A. P. C., & Hanson, B. J. (2011). Comparison of the efficiency of antibody selection from semi-synthetic scFv and non-immune Fab phage display libraries against protein targets for rapid development of diagnostic immunoassays. Journal of Immunological Methods, 373(1-2), 79–88. doi:10.1016/j.jim.2011.08.005

Chattopadhyay, P. K., Gierahn, T. M., Roederer, M., & Love, J. C. (2014). Single-cell technologies for monitoring immune systems. Nature Immunology, 15(2), 128–35. doi:10.1038/ni.2796

Chen, G., & Sidhu, S. S. (2014). Design and generation of synthetic antibody libraries for phage display. Methods in Molecular Biology (Clifton, N.J.), 1131, 113–31. doi:10.1007/978-1-62703-992-5_8

De Bruyn, M., Bremer, E., & Helfrich, W. (2013). Antibody-based fusion proteins to target death receptors in cancer. Cancer Letters, 332(2), 175–83. doi:10.1016/j.canlet.2010.11.006

Debs, B. E., Utharala, R., Balyasnikova, I. V., Griffiths, A. D., & Merten, C. A. (2012). Functional single-cell hybridoma screening using droplet-based microfluidics. Proceedings of the National Academy of Sciences. doi:10.1073/pnas.1204514109

Demarest, S. J., & Glaser, S. M. (2008). Antibody therapeutics, antibody engineering, and the merits of protein stability. Current Opinion in Drug Discovery & Development, 11(5), 675–87. Retrieved from

Dreher, M. L., Gherardi, E., Skerra, A., & Milstein, C. (1991). Colony assays for antibody fragments expressed in bacteria. Journal of Immunological Methods, 139(2), 197–205. Retrieved from

Dübel, S. (2007). Recombinant therapeutic antibodies. Applied Microbiology and Biotechnology, 74(4), 723–9. doi:10.1007/s00253-006-0810-y

Feldhaus, M. J., & Siegel, R. W. (2004). Yeast display of antibody fragments: a discovery and characterization platform. Journal of Immunological Methods, 290(1-2), 69–80. doi:10.1016/j.jim.2004.04.009

Geyer, C. R., McCafferty, J., Dübel, S., Bradbury, A. R. M., & Sidhu, S. S. (2012). Recombinant antibodies and in vitro selection technologies. Methods in Molecular Biology (Clifton, N.J.), 901, 11–32. doi:10.1007/978-1-61779-931-0_2

Giordano, R. J., Cardó-Vila, M., Lahdenranta, J., Pasqualini, R., & Arap, W. (2001). Biopanning and rapid analysis of selective interactive ligands. Nature Medicine, 7(11), 1249–53. doi:10.1038/nm1101-1249

Giovannoni, L., Viti, F., Zardi, L., & Neri, D. (2001). Isolation of anti-angiogenesis antibodies from a large combinatorial repertoire by colony filter screening. Nucleic Acids Research, 29(5), E27. Retrieved from

Griffiths, A. D., Williams, S. C., Hartley, O., Tomlinson, I. M., Waterhouse, P., Crosby, W. L., … Allison, T. J. (1994). Isolation of high affinity human antibodies directly from large synthetic repertoires. The EMBO Journal, 13(14), 3245–60. Retrieved from

Hagemeyer, C. E., von Zur Muhlen, C., von Elverfeldt, D., & Peter, K. (2009). Single-chain antibodies as diagnostic tools and therapeutic agents. Thrombosis and Haemostasis, 101(6), 1012–9. Retrieved from

Haque, A., & Tonks, N. K. (2012). The use of phage display to generate conformation-sensor recombinant antibodies. Nature Protocols, 7(12), 2127–43. doi:10.1038/nprot.2012.132

Holliger, P., & Hudson, P. J. (2005). Engineered antibody fragments and the rise of single domains. Nature Biotechnology, 23(9), 1126–36. doi:10.1038/nbt1142

Hoogenboom, H. R. (2005). Selecting and screening recombinant antibody libraries. Nature Biotechnology, 23(9), 1105–16. doi:10.1038/nbt1126

Hust, M., Frenzel, A., Schirrmann, T., & Dübel, S. (2014). Selection of recombinant antibodies from antibody gene libraries. Methods in Molecular Biology (Clifton, N.J.), 1101, 305–20. doi:10.1007/978-1-62703-721-1_14

Jäger, V., Büssow, K., Wagner, A., Weber, S., Hust, M., Frenzel, A., & Schirrmann, T. (2013). High level transient production of recombinant antibodies and antibody fusion proteins in HEK293 cells. BMC Biotechnology, 13, 52. doi:10.1186/1472-6750-13-52

Jostock, T., & Dübel, S. (2005). Screening of molecular repertoires by microbial surface display. Combinatorial Chemistry & High Throughput Screening, 8(2), 127–33. Retrieved from

Kaiser, P. D., Maier, J., Traenkle, B., Emele, F., & Rothbauer, U. (2014). Recent progress in generating intracellular functional antibody fragments to target and trace cellular components in living cells. Biochimica et Biophysica Acta, 1844(11), 1933–1942. doi:10.1016/j.bbapap.2014.04.019

Kato, M., & Hanyu, Y. (2013). Construction of an scFv library by enzymatic assembly of VL and VH genes. Journal of Immunological Methods, 396(1-2), 15–22.

Kato, M., & Hanyu, Y. (2015). Direct cloning from antibody libraries. to be submitted to Journal of Immunological Methods.

Kato, M., Sasamori, E., Chiba, T., & Hanyu, Y. (2011). Cell activation by CpG ODN leads to improved electrofusion in hybridoma production. Journal of Immunological Methods, 373(1-2), 102–110.

Köhler, G., & Milstein, C. (1975). Continuous cultures of fused cells secreting antibody of predefined specificity. Nature, 256(5517), 495–7. Retrieved from

Kontermann, R. E. (2010). Alternative antibody formats. Current Opinion in Molecular Therapeutics, 12(2), 176–83. Retrieved from

McCafferty, J., Griffiths, A. D., Winter, G., & Chiswell, D. J. (1990). Phage antibodies: filamentous phage displaying antibody variable domains. Nature, 348(6301), 552–4. doi:10.1038/348552a0

Miethe, S., Meyer, T., Wöhl-Bruhn, S., Frenzel, A., Schirrmann, T., Dübel, S., & Hust, M. (2013). Production of single chain fragment variable (scFv) antibodies in Escherichia coli using the LEXTM bioreactor. Journal of Biotechnology, 163(2), 105–11. doi:10.1016/j.jbiotec.2012.07.011

Morino, K., Katsumi, H., Akahori, Y., Iba, Y., Shinohara, M., Ukai, Y., … Kurosawa, Y. (2001). Antibody fusions with fluorescent proteins: a versatile reagent for profiling protein expression. Journal of Immunological Methods, 257(1-2), 175–84. Retrieved from

Müller, D. (2010). scFv by Two-Step Cloning. In R. Kontermann & S. Dübel (Eds.), Antibody Engineering (pp. 55–59). Springer Berlin Heidelberg.

Nelson, A. L. (2010). Antibody fragments: Hope and hype. mAbs. doi:10.4161/mabs.2.1.10786

Nettleship, J. E., Ren, J., Rahman, N., Berrow, N. S., Hatherley, D., Barclay, a N., & Owens, R. J. (2008). A pipeline for the production of antibody fragments for structural studies using transient expression in HEK 293T cells. Protein Expression and Purification, 62(1), 83–9. doi:10.1016/j.pep.2008.06.017

Parmley, S. F., & Smith, G. P. (1988). Antibody-selectable filamentous fd phage vectors: affinity purification of target genes. Gene, 73(2), 305–18. Retrieved from

Persson, M. A., Caothien, R. H., & Burton, D. R. (1991). Generation of diverse high-affinity human monoclonal antibodies by repertoire cloning. Proceedings of the National Academy of Sciences of the United States of America, 88(6), 2432–6. Retrieved from

Prassler, J., Steidl, S., & Urlinger, S. (2009). In vitro affinity maturation of HuCAL antibodies: complementarity determining region exchange and RapMAT technology. Immunotherapy, 1(4), 571–83. doi:10.2217/imt.09.23

Prassler, J., Thiel, S., Pracht, C., Polzer, A., Peters, S., Bauer, M., … Enzelberger, M. (2011). HuCAL PLATINUM, a Synthetic Fab Library Optimized for Sequence Diversity and Superior Performance in Mammalian Expression Systems. Journal of Molecular Biology, 413(1), 261–278. doi:10.1016/j.jmb.2011.08.012

Rauth, S., Schlapschy, M., & Skerra, A. (2010). Selection of Antibody Fragments by Means of the Filter-Sandwich Colony Screening Assay. In R. Kontermann & S. Dübel (Eds.), Antibody Engineering (pp. 255–266). Springer Berlin Heidelberg.

Reichert, J. M. (2015). Antibodies to watch in 2015. mAbs, 7(1), 1–8. doi:10.4161/19420862.2015.988944

Rothe, C., Urlinger, S., Löhning, C., Prassler, J., Stark, Y., Jäger, U., … Urban, M. (2008). The human combinatorial antibody library HuCAL GOLD combines diversification of all six CDRs according to the natural immune system with a novel display method for efficient selection of high-affinity antibodies. Journal of Molecular Biology, 376(4), 1182–200. doi:10.1016/j.jmb.2007.12.018

Schaefer, J. V., Honegger, A., & Plückthun, A. (2010). Construction of scFv Fragments from Hybridoma or Spleen Cells by PCR Assembly. In R. Kontermann & S. Dübel (Eds.), Antibody Engineering (pp. 21–44). Springer Berlin Heidelberg.

Schaefer, J. V., Lindner, P., & Plückthun, A. (2010). Miniantibodies. In R. Kontermann & S. Dübel (Eds.), Antibody Engineering (pp. 85–99). Springer Berlin Heidelberg.

Schaffitzel, C., Hanes, J., Jermutus, L., & Plückthun, A. (1999). Ribosome display: an in vitro method for selection and evolution of antibodies from libraries. Journal of Immunological Methods, 231(1-2), 119–35. Retrieved from

Schirrmann, T., Al-Halabi, L., Dübel, S., & Hust, M. (2008). Production systems for recombinant antibodies. Frontiers in Bioscience : A Journal and Virtual Library, 13, 4576–94. Retrieved from

Schirrmann, T., Meyer, T., Schütte, M., Frenzel, A., & Hust, M. (2011). Phage display for the generation of antibodies for proteome research, diagnostics and therapy. Molecules (Basel, Switzerland), 16(1), 412–26. doi:10.3390/molecules16010412

Schofield, D. J., Pope, A. R., Clementel, V., Buckell, J., Chapple, S. D., Clarke, K. F., … McCafferty, J. (2007). Application of phage display to high throughput antibody generation and characterization. Genome Biology, 8(11), R254. doi:10.1186/gb-2007-8-11-r254

Skerra, A., Dreher, M. L., & Winter, G. (1991). Filter screening of antibody Fab fragments secreted from individual bacterial colonies: specific detection of antigen binding with a two-membrane system. Analytical Biochemistry, 196(1), 151–5. Retrieved from

Steinwand, M., Droste, P., Frenzel, A., Hust, M., Dübel, S., & Schirrmann, T. (2014). The influence of antibody fragment format on phage display based affinity maturation of IgG. mAbs, 6(1), 204–18. doi:10.4161/mabs.27227

Thie, H., Voedisch, B., Dübel, S., Hust, M., & Schirrmann, T. (2009). Affinity maturation by phage display. Methods in Molecular Biology (Clifton, N.J.), 525, 309–22, xv. doi:10.1007/978-1-59745-554-1_16

Weiner, G. J. (2015). Building better monoclonal antibody-based therapeutics. Nature Reviews Cancer, 15(6), 361–370. doi:10.1038/nrc3930

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