Development of an immunocytochemical assay to detect human CFTR expression following gene therapy

H. Davidson, A. Wilson, R. D. Gray, A. Horsley, I. Pringle, G. McLachlan, J. Gibson, E. Holder, L. Jones, A. Doherty, R. Coles, S. Sumner-Jones, M. Wasowicz, M. Manvell, U. Griesenbach, S. C. Hyde, D. R. Gill, J. Davies, D. D. S. Collie, E. AltonD. J. Porteous, A. C. Boyd

Research output: Contribution to journalMeeting abstractpeer-review


To assess gene therapy treatment for cystic fibrosis (CF) in clinical trials it is essential to develop robust assays that can accurately detect transgene expression in human airway epithelial cells. Our aim was to develop a reproducible immunocytochemical assay for human CFTR protein which can measure both endogenous CFTR levels and augmented CFTR expression after gene delivery.
We characterised an antibody (G449) which satisfied the criteria for use in clinical trials. We optimised our immunocytochemistry method and identified G449 dilutions at which endogenous CFTR levels were negligible in CF samples, thus enhancing detection of transgenic CFTR protein. After developing a transfection technique for brushed human nasal epithelial cells, we transfected non-CF and CF cells with a clinically relevant CpG-free plasmid encoding human CFTR.
The optimised immunocytochemistry method gave improved discrimination between CF and non-CF samples. Transfection of a CFTR expression vector into primary nasal epithelial cells resulted in detectable RNA and protein expression. CFTR protein was present in 0.05-10% of non-CF cells and 0.02-0.8% of CF cells.
We have developed a sensitive, clinically relevant immunocytochemical assay for CFTR protein and have used it to detect transgene-expressed CFTR in transfected human primary airway epithelial cells
Original languageEnglish
Pages (from-to)416
Number of pages1
JournalHuman Gene Therapy
Issue number4
Publication statusPublished - 2008
EventThe 5th Annual BSGT Annual Conference - Heriot-Watt University, Edinburgh, United Kingdom
Duration: 7 Apr 20089 Apr 2008


  • Cells, Cultured
  • Cystic Fibrosis
  • Cystic Fibrosis Transmembrane Conductance Regulator
  • Epithelial Cells
  • Genetic Therapy
  • Green fluorescent protein
  • Immunohistochemistry
  • Nasal Mucosa
  • Reproducibility of Results
  • Respiratory Mucosa
  • Transgenes


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