Immunogenomic engineering of a plug-and-(dis)play hybridoma platform

Mark Pogson, Cristina Parola, William J. Kelton, Paul Heuberger and Sai T. Reddy ()
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Mark Pogson: ETH Zurich
Cristina Parola: ETH Zurich
William J. Kelton: ETH Zurich
Paul Heuberger: ETH Zurich
Sai T. Reddy: ETH Zurich

Nature Communications, 2016, vol. 7, issue 1, 1-10

Abstract: Abstract Hybridomas, fusions of primary mouse B cells and myelomas, are stable, rapidly-proliferating cell lines widely utilized for antibody screening and production. Antibody specificity of a hybridoma clone is determined by the immunoglobulin sequence of the primary B cell. Here we report a platform for rapid reprogramming of hybridoma antibody specificity by immunogenomic engineering. Here we use CRISPR-Cas9 to generate double-stranded breaks in immunoglobulin loci, enabling deletion of the native variable light chain and replacement of the endogenous variable heavy chain with a fluorescent reporter protein (mRuby). New antibody genes are introduced by Cas9-targeting of mRuby for replacement with a donor construct encoding a light chain and a variable heavy chain, resulting in full-length antibody expression. Since hybridomas surface express and secrete antibodies, reprogrammed cells are isolated using flow cytometry and cell culture supernatant is used for antibody production. Plug-and-(dis)play hybridomas can be reprogrammed with only a single transfection and screening step.

Date: 2016
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Persistent link: https://EconPapers.repec.org/RePEc:nat:natcom:v:7:y:2016:i:1:d:10.1038_ncomms12535

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DOI: 10.1038/ncomms12535

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