SCIENCE@home

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

M. Pogson,Cristina Parola,William J Kelton,P. Heuberger,S. Reddy

Published 2016 in Nature Communications

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. Hybridomas are widely used for antibody screening and production due to their genetic stability and rapid proliferation. Here the authors demonstrate the rapid reprogramming of antibody specificity in hybridomas using CRISPR-Cas9.

PUBLICATION RECORD

CITATION MAP

EXTRACTION MAP

CLAIMS

  • No claims are published for this paper.

CONCEPTS

  • No concepts are published for this paper.

REFERENCES

Showing 1-68 of 68 references · Page 1 of 1

CITED BY

Showing 1-52 of 52 citing papers · Page 1 of 1