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Detecting Cytosolic Peptide Delivery with the GFP Complementation Assay in the Low Micromolar Range |
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| Authors: | Samuel Schmidt Dr. Merel J. W. Adjobo‐Hermans Dr. Rike Wallbrecher Dr. Wouter P. R. Verdurmen Petra H. M. Bovée‐Geurts Jenny van Oostrum Dr. Francesca Milletti Dr. Thilo Enderle Prof. Dr. Roland Brock |
| Affiliation: | 1. Department of Biochemistry (286), Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Geert Grooteplein 28, 6525 GA Nijmegen (The Netherlands);2. Present address: Department of Biochemistry, University of Zurich, Winterthurerstrasse 190, 8057 Zurich (Switzerland);3. Roche Pharmaceutical Research and Early Development, Roche Innovation Center New York, 430 East 29th Street, 10016 New York, NY (USA);4. Roche Pharmaceutical Research and Early Development, Roche Innovation Center Basel, F. Hoffmann‐La Roche Ltd, Grenzacherstrasse 124, 4070 Basel (Switzerland) |
| Abstract: | Transfection of cells with a plasmid encoding for the first ten strands of the GFP protein (GFP1‐10) provides the means to detect cytosolic peptide import at low micromolar concentrations. Cytosolic import of the eleventh strand of the GFP protein either by electroporation or by cell‐penetrating peptide‐mediated import leads to formation of the full‐length GFP protein and fluorescence. An increase in sensitivity is achieved through structural modifications of the peptide and the expression of GFP1‐10 as a fusion protein with mCherry. |
| Keywords: | cell uptake cell‐penetrating peptides drug delivery fragment complementation assays lipid membranes |