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Nat Commun. 2016 Aug 17;7:12535. doi: 10.1038/ncomms12535.

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

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Department of Biosystems Science and Engineering, ETH Zurich, Mattenstrasse 26, Basel 4058, Switzerland.
Life Science Zurich Graduate School, Systems Biology, ETH Zurich, University of Zurich, Zurich 8057, Switzerland.


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.

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