Homologous Recombination in Hybridoma Cells: Heavy Chain Chimeric Antibody Produced by Gene Targeting

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 86; no. 21; pp. 8507 - 8511
• Main Authors: Fell, H. Perry, Yarnold, Susan, Hellström, I., Hellström, K. E., Folger, Kim R.
• Format: Journal Article
• Language: English
• Published: Washington, DC National Academy of Sciences of the United States of America 01.11.1989; National Acad Sciences
• Subjects: Animals; Antibodies; Antibody-Dependent Cell Cytotoxicity; Bacteriophages; Biological and medical sciences; Blotting, Western; Cell Line; Cell lines; Chimera; DNA; Enzyme-Linked Immunosorbent Assay; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Genes, Immunoglobulin; Genetic loci; Homologous recombination; Humans; Hybridomas; Hybridomas - immunology; Immunoglobulin G - genetics; Immunoglobulin Heavy Chains - genetics; Mice; Molecular immunology; Molecules; Plasmacytoma; Plasmids; Recombination, Genetic; Restriction Mapping; Techniques; Transfection; Human; Recombination; Antibody; heavy-Peptide chain; Rodentia; Hybridoma; Vertebrata; Mammalia; Cell line; Transfection; Gene; Mouse; DNA; Antibody-dependent cell cytotoxicity; Immunoblotting assay; ELISA assay; Chimera
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.86.21.8507

We demonstrate that murine myeloma cells can efficiently mediate homologous recombination. The murine myeloma cell line J558L was shown to appropriately recombine two transfected DNA molecules in ≈ 30% of cells that received and integrated intact copies of both molecules. This activity was then exploited to direct major reconstructions of an endogenous locus within a hybridoma cell line. Production of antigen-specific chimeric heavy chain was achieved by targeting the human IgG1 heavy chain constant region (Cγ 1) exons to the genomic heavy chain locus of a hybridoma cell line secreting antibody specific for a human tumor-associated antigen. The frequency of productive genomic recombinations was ≈ 1 in 200 transfectants, with accumulation of the chimeric protein reaching >20 μ g/ml in culture supernatants.