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Cloning and Reexpression of a Functional Human IgM Anti-Lipid A Antibody

The human hybridoma cell line HR78 secretes a human antibody of the IgM isotype directed against bacterial endotoxin. The cell line produces low levels of antibody and, more importantly, the antibody product is likely to be impure since two and perhaps more species of heavy chain are being synthesized as judged by cloning and expression studies. To address these issues, the cell line was used as a source of mRNA for the construction of cDNA libraries and the subsequent isolation of the sequences encoding heavy and light chains. Expression of these chains in a non-immunoglobulin producing murine hybridoma cell line resulted in a monoclonal antibody that bound antigen in a manner essentially identical to that observed for the parental antibody. Moreover, the level of expression of human IgM in selected clones was increased approximately 50-fold over the parental cell line and was comparable to or better than antibody expression by a typical murine hybridoma. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Hybridoma Mary Ann Liebert

Cloning and Reexpression of a Functional Human IgM Anti-Lipid A Antibody

Abstract

The human hybridoma cell line HR78 secretes a human antibody of the IgM isotype directed against bacterial endotoxin. The cell line produces low levels of antibody and, more importantly, the antibody product is likely to be impure since two and perhaps more species of heavy chain are being synthesized as judged by cloning and expression studies. To address these issues, the cell line was used as a source of mRNA for the construction of cDNA libraries and the subsequent isolation of the sequences encoding heavy and light chains. Expression of these chains in a non-immunoglobulin producing murine hybridoma cell line resulted in a monoclonal antibody that bound antigen in a manner essentially identical to that observed for the parental antibody. Moreover, the level of expression of human IgM in selected clones was increased approximately 50-fold over the parental cell line and was comparable to or better than antibody expression by a typical murine hybridoma.
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