A rapid screening and production method using a novel mammalian cell display to isolate human monoclonal antibodies
| Autor: | Tomoya Kinjo, Sanetaka Shirahata, Shin Ei Matsumoto, Takeki Hamasaki, Kiichiro Teruya, Hayato Tanaka, Yoshinori Katakura, Saiko Kazuno, Ken Ichi Kusumoto, Kosuke Tomimatsu, Makiko Yamashita, Hidekazu Nakanishi, Shigeru Kabayama |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
medicine.drug_class
Molecular Sequence Data Cell Gene Conversion Biophysics CHO Cells Biology Monoclonal antibody Biochemistry Affinity maturation Cricetulus Antigen Peptide Library Cricetinae medicine Animals Humans Amino Acid Sequence Transgenes Antibody-Producing Cells Molecular Biology Recombination Genetic Receptors IgE Chinese hamster ovary cell Cell Membrane Antibodies Monoclonal Cell Biology Transfection Molecular biology medicine.anatomical_structure Immunoglobulin G Monoclonal biology.protein Antibody Cell Surface Display Techniques |
| Zdroj: | Biochemical and Biophysical Research Communications. 441:59-64 |
| ISSN: | 0006-291X |
| Popis: | Antibody display methods are increasingly being used to produce human monoclonal antibodies for disease therapy. Rapid screening and isolation of specific human antibody genes are valuable for producing human monoclonal antibodies showing high specificity and affinity. In this report, we describe a novel mammalian cell display method in which whole human IgG is displayed on the cell surface of CHO cells. Cells expressing antigen-specific human monoclonal IgGs with high affinity on the cell surface after normal folding and posttranscriptional modification were screened using a cell sorter. The membrane-type IgG-expressing CHO cells were then converted to IgG-secreting cells by transfection with a plasmid coding Cre recombinase. This mammalian cell display method was applied to in vitro affinity maturation of monoclonal C9 IgG specific to the human high-affinity IgE receptor (FcεRIα). The CDR3 of the C9 heavy chain variable region gene was randomly mutated and inserted into pcDNA5FRT/IgG. A C9 IgG (CDRH3r)-expressing CHO cell display library consisting of 1.1×10(6) independent clones was constructed. IgG-displaying cells showing high reactivity to FcεRIα antigen were screened by the cell sorter, resulting in the establishment of a CHO cell line producing with higher reactivity than the parent C9 IgG. |
| Databáze: | OpenAIRE |
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