On chip electrofusion of single human B cells and mouse myeloma cells for efficient hybridoma generation

Evelien Kemna, F. Wolbers, Albert van den Berg, I. Vermes

    Research output: Contribution to journalArticleAcademicpeer-review

    27 Citations (Scopus)

    Abstract

    This article describes the development and full characterization of a microfluidic chip for electrofusion of human peripheral blood B-cells and mouse myeloma (NS-1) cells to generate hybridomas. The chip consists of an array of 783 traps, with dimensions that were optimized to obtain a final cell pairing efficiency of 3376%. B cells were stained with a cytoplasmic stain CFDA to assess the different stages of cell fusion, i.e. dye transfer to NS-1 cells (initiating fusion) and membrane reorganization (advanced fusion). Six DC pulses of 100 ms (2.5 kV/cm) combined with an AC field (30 s, 2 MHz, 500 V/cm) and pronase treatment resulted in the highest electrofusion efficiency of paired cells (51711%). Hybridoma formation, with a yield of 0.33 and 1.2%, was observed after culturing the fused cells for 14 days in conditioned medium. This work provides valuable leads to improve the current electrofusion protocols for the production of human antibodies for diagnostic and therapeutic applications.
    Original languageUndefined
    Pages (from-to)3138-3146
    Number of pages9
    JournalElectrophoresis
    Volume32
    Issue number22
    DOIs
    Publication statusPublished - Nov 2011

    Keywords

    • IR-78991
    • Electrofusion / Human B cells / Hybridoma culture / Microfluidics / Mousemyeloma cells
    • EWI-21016
    • METIS-281651

    Cite this

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    title = "On chip electrofusion of single human B cells and mouse myeloma cells for efficient hybridoma generation",
    abstract = "This article describes the development and full characterization of a microfluidic chip for electrofusion of human peripheral blood B-cells and mouse myeloma (NS-1) cells to generate hybridomas. The chip consists of an array of 783 traps, with dimensions that were optimized to obtain a final cell pairing efficiency of 3376{\%}. B cells were stained with a cytoplasmic stain CFDA to assess the different stages of cell fusion, i.e. dye transfer to NS-1 cells (initiating fusion) and membrane reorganization (advanced fusion). Six DC pulses of 100 ms (2.5 kV/cm) combined with an AC field (30 s, 2 MHz, 500 V/cm) and pronase treatment resulted in the highest electrofusion efficiency of paired cells (51711{\%}). Hybridoma formation, with a yield of 0.33 and 1.2{\%}, was observed after culturing the fused cells for 14 days in conditioned medium. This work provides valuable leads to improve the current electrofusion protocols for the production of human antibodies for diagnostic and therapeutic applications.",
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    author = "Evelien Kemna and F. Wolbers and {van den Berg}, Albert and I. Vermes",
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    doi = "10.1002/elps.201100227",
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    volume = "32",
    pages = "3138--3146",
    journal = "Electrophoresis",
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    On chip electrofusion of single human B cells and mouse myeloma cells for efficient hybridoma generation. / Kemna, Evelien; Wolbers, F.; van den Berg, Albert; Vermes, I.

    In: Electrophoresis, Vol. 32, No. 22, 11.2011, p. 3138-3146.

    Research output: Contribution to journalArticleAcademicpeer-review

    TY - JOUR

    T1 - On chip electrofusion of single human B cells and mouse myeloma cells for efficient hybridoma generation

    AU - Kemna, Evelien

    AU - Wolbers, F.

    AU - van den Berg, Albert

    AU - Vermes, I.

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    PY - 2011/11

    Y1 - 2011/11

    N2 - This article describes the development and full characterization of a microfluidic chip for electrofusion of human peripheral blood B-cells and mouse myeloma (NS-1) cells to generate hybridomas. The chip consists of an array of 783 traps, with dimensions that were optimized to obtain a final cell pairing efficiency of 3376%. B cells were stained with a cytoplasmic stain CFDA to assess the different stages of cell fusion, i.e. dye transfer to NS-1 cells (initiating fusion) and membrane reorganization (advanced fusion). Six DC pulses of 100 ms (2.5 kV/cm) combined with an AC field (30 s, 2 MHz, 500 V/cm) and pronase treatment resulted in the highest electrofusion efficiency of paired cells (51711%). Hybridoma formation, with a yield of 0.33 and 1.2%, was observed after culturing the fused cells for 14 days in conditioned medium. This work provides valuable leads to improve the current electrofusion protocols for the production of human antibodies for diagnostic and therapeutic applications.

    AB - This article describes the development and full characterization of a microfluidic chip for electrofusion of human peripheral blood B-cells and mouse myeloma (NS-1) cells to generate hybridomas. The chip consists of an array of 783 traps, with dimensions that were optimized to obtain a final cell pairing efficiency of 3376%. B cells were stained with a cytoplasmic stain CFDA to assess the different stages of cell fusion, i.e. dye transfer to NS-1 cells (initiating fusion) and membrane reorganization (advanced fusion). Six DC pulses of 100 ms (2.5 kV/cm) combined with an AC field (30 s, 2 MHz, 500 V/cm) and pronase treatment resulted in the highest electrofusion efficiency of paired cells (51711%). Hybridoma formation, with a yield of 0.33 and 1.2%, was observed after culturing the fused cells for 14 days in conditioned medium. This work provides valuable leads to improve the current electrofusion protocols for the production of human antibodies for diagnostic and therapeutic applications.

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