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

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

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.

Original languageEnglish
Pages (from-to)1069-1083
Number of pages15
JournalmAbs
Volume6
Issue number4
DOIs
Publication statusPublished - 2014

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Keywords

  • Activation-induced cytidine affinity maturation
  • AID
  • Anti-PEG
  • Class switch recombination
  • Germinal center reaction
  • Hybridoma
  • PEG
  • Polyethylene glycol
  • Somatic hypermutation
  • Temperature-dependent binding
  • Temperature-selective antibody

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Medicine(all)

Cite this

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