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Sci Rep. 2014 Jul 30;4:5885. doi: 10.1038/srep05885.

A tool kit for rapid cloning and expression of recombinant antibodies.

Author information

1
1] NIHR Biomedical Research Centre at Guy's and St. Thomas's Hospitals and King's College London, London, UK [2] Randall Division of Cell and Molecular Biophysics, King's College London, Guy's Campus, London SE1 1UL, UK.
2
1] NIHR Biomedical Research Centre at Guy's and St. Thomas's Hospitals and King's College London, London, UK [2] St. John's Institute of Dermatology, Division of Genetics and Molecular Medicine, King's College London School of Medicine, King's College London, London SE1 9RT, UK.
3
1] NIHR Biomedical Research Centre at Guy's and St. Thomas's Hospitals and King's College London, London, UK [2] St. John's Institute of Dermatology, Division of Genetics and Molecular Medicine, King's College London School of Medicine, King's College London, London SE1 9RT, UK [3] Division of Cancer Studies, King's College London, 3rd Floor Bermondsey Wing, Guy's Hospital, London SE1 9RT, UK.
4
Randall Division of Cell and Molecular Biophysics, King's College London, Guy's Campus, London SE1 1UL, UK.

Abstract

Over the last four decades, molecular cloning has evolved tremendously. Efficient products allowing assembly of multiple DNA fragments have become available. However, cost-effective tools for engineering antibodies of different specificities, isotypes and species are still needed for many research and clinical applications in academia. Here, we report a method for one-step assembly of antibody heavy- and light-chain DNAs into a single mammalian expression vector, starting from DNAs encoding the desired variable and constant regions, which allows antibodies of different isotypes and specificity to be rapidly generated. As a proof of principle we have cloned, expressed and characterized functional recombinant tumor-associated antigen-specific chimeric IgE/κ and IgG1/κ, as well as recombinant grass pollen allergen Phl p 7 specific fully human IgE/λ and IgG4/λ antibodies. This method utilizing the antibody expression vectors, available at Addgene, has many applications, including the potential to support simultaneous processing of antibody panels, to facilitate mechanistic studies of antigen-antibody interactions and to conduct early evaluations of antibody functions.

PMID:
25073855
PMCID:
PMC4115235
DOI:
10.1038/srep05885
[Indexed for MEDLINE]
Free PMC Article

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