Mimicking the germinal center reaction in hybridoma cells to isolate temperature-selective anti-PEG antibodies

Yu Cheng Su, Talal S. Al-Qaisi, Hsin Yi Tung, Tian Lu Cheng, Kuo Hsiang Chuang, Bing Mae Chen, Steve R. Roffler

研究成果: 雜誌貢獻文章

6 引文 斯高帕斯(Scopus)

摘要

Modification of antibody class and binding properties typically requires cloning of antibody genes, antibody library construction, phage or yeast display and recombinant antibody expression. Here, we describe an alternative "cloning-free" approach to generate antibodies with altered antigen-binding and heavy chain isotype by mimicking the germinal center reaction in antibody-secreting hybridoma cells. This was accomplished by lentiviral transduction and controllable expression of activation-induced cytidine deaminase (AID) to generate somatic hypermutation and class switch recombination in antibody genes coupled with high-throughput fluorescence-activated cell sorting (FACS) of hybridoma cells to detect altered antibody binding properties. Starting from a single established hybridoma clone, we isolated mutated antibodies that bind to a low-temperature structure of polyethylene glycol (PEG), a polymer widely used in nanotechnology, biotechnology and pharmaceuticals. FACS of AID-infected hybridoma cells also facilitated rapid identification of class switched variants of monoclonal IgM to monoclonal IgG. Mimicking the germinal center reaction in hybridoma cells may offer a general method to identify and isolate antibodies with altered binding properties and class-switched heavy chains without the need to carry out DNA library construction, antibody engineering and recombinant protein expression.

原文英語
頁(從 - 到)1069-1083
頁數15
期刊mAbs
6
發行號4
DOIs
出版狀態已發佈 - 2014

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Medicine(all)

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  • 引用此

    Su, Y. C., Al-Qaisi, T. S., Tung, H. Y., Cheng, T. L., Chuang, K. H., Chen, B. M., & Roffler, S. R. (2014). Mimicking the germinal center reaction in hybridoma cells to isolate temperature-selective anti-PEG antibodies. mAbs, 6(4), 1069-1083. https://doi.org/10.4161/mabs.29124