Cloning, Expression, and Purification of Monoclonal Antibodies in scFv-Fc Format

  • Jiahui Yang
  • Christoph RaderEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 901)


This protocol describes the generation of monoclonal antibodies in single-chain variable fragment (scFv)-Fc format. It includes the cloning of the scFv-Fc expression cassette into a mammalian expression vector followed by transient transfection of mammalian cells and purification by protein A affinity chromatography. The protocol is intended for applications in basic and preclinical research that require rapid access to milligram amounts of protein.

Key words

Monoclonal antibodies scFv-Fc format Hemagglutinin tag FreeStyle 293F cells Protein A affinity chromatography 



This work was supported by the Intramural Research Program of the Center for Cancer Research, National Cancer Institute, National Institutes of Health. We thank Marko Modric for comments.


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Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  1. 1.Experimental Transplantation and Immunology BranchCenter for Cancer Research, National Cancer Institute, National Institutes of HealthBethesdaUSA
  2. 2.Department of Cancer Biology and Department of Molecular TherapeuticsThe Scripps Research InstituteScripps Florida JupiterUSA

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