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Expression Cloning of Antibodies from Single Human B Cells

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Lymphoma

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1956))

Abstract

The majority of lymphomas originate from B cells at the germinal center stage. Preferential selection of B-cell clones by a limited set of antigens has been suggested to drive lymphoma development. While recent studies in chronic lymphocytic leukemia have shown that self-reactive B-cell receptors (BCR) can generate cell-autonomous signaling and proliferation, our knowledge about the role of BCRs for the development or survival of other lymphomas remains limited. Here, we describe a strategy to characterize the antibody reactivity of human B cells. The approach allows the unbiased characterization of the human antibody repertoire at single-cell level through the generation of recombinant monoclonal antibodies from single primary human B cells of defined origin. This protocol offers a detailed description of the method starting from the flow-cytometric isolation of single human B cells to the reverse transcription-polymerase chain reaction (RT-PCR)-based amplification of the expressed immunoglobulin (Ig) transcripts (IGH, IGK, and IGL) and their subsequent cloning into expression vectors for the in vitro production of recombinant monoclonal antibodies. The strategy may be used to obtain information on the clonal evolution of B-cell lymphomas by single-cell sequencing of Ig transcripts and on the antibody reactivity of human lymphoma B cells.

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Authors and Affiliations

  1. Division of B-Cell Immunology, German Cancer Research Center (DKFZ), Heidelberg, Germany

    Hedda Wardemann & Christian E. Busse

Authors
  1. Hedda Wardemann
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  2. Christian E. Busse
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Correspondence to Hedda Wardemann .

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Editors and Affiliations

  1. Institute of Cell Biology (Cancer Research), Medical School, University of Duisburg-Essen, Nordrhein-Westfalen, Essen, Germany

    Ralf Küppers

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Wardemann, H., Busse, C.E. (2019). Expression Cloning of Antibodies from Single Human B Cells. In: Küppers, R. (eds) Lymphoma. Methods in Molecular Biology, vol 1956. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-9151-8_5

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  • DOI: https://doi.org/10.1007/978-1-4939-9151-8_5

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-9150-1

  • Online ISBN: 978-1-4939-9151-8

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