Isolation of full-length IgG antibodies from combinatorial libraries expressed in the cytoplasm of Escherichia coli

We describe a facile and robust genetic selection for isolating full-length IgG antibodies from combinatorial libraries expressed in the cytoplasm of the genetically engineered Escherichia coli strain, SHuffle. The method is based on the transport of a bifunctional substrate comprised of an antigen fused to chloramphenicol acetyltransferase, which allows positive selection of bacterial cells co-expressing cytoplasmic IgGs called ‘cyclonals’ that specifically capture the chimeric antigen and sequester the antibiotic resistance marker in the cytoplasm. The selective power of this approach was demonstrated by facile isolation of novel complementarity-determining regions for a cyclonal that specifically recognized the basic-region leucine zipper domain of the yeast transcriptional activator protein Gcn4.Competing Interest StatementM.P.D. has ownership interest (including stock, patents, etc.) in SwiftScale Biologics, Inc. M.P.D.’s interests are reviewed and managed by Cornell University in accordance with their conflict of interest policies. M.B. is employed by NEB, which commercializes SHuffle cells. All other authors declare no competing interests.View Full Text