Kulkarni, Rajan P. and Mishra, Swaroop and Fraser, Scott E. and Davis, Mark E. (2005) Single Cell Kinetics of Intracellular, Nonviral, Nucleic Acid Delivery Vehicle Acidification and Trafficking. Bioconjugate Chemistry, 16 (4). pp. 986-994. ISSN 1043-1802. doi:10.1021/bc050081u. https://resolver.caltech.edu/CaltechAUTHORS:20160226-114842424
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Abstract
Mechanistic understanding of the intracellular trafficking of nonviral nucleic acid delivery vehicles remains elusive. A live, single cell-based assay is described here that is used to investigate and quantitate the spatiotemporal, intracellular pH microenvironment of polymeric-based nucleic acid delivery vehicles. Polycations such as polyethylenimine (PEI), poly-L-lysine (PLL), β-cyclodextrin-containing polymers lacking or possessing imidazole termini (CDP or CDP-imid), and cyclodextrin-grafted PEI (CD-PEI) are used to deliver an oligonucleotide containing a single fluorophore with two emission lines that can be employed to measure the pH. Delivery vehicles were also sterically stabilized by addition of poly(ethylene glycol) (PEG) and investigated. The intracellular trafficking data obtained via this new methodology show that vectors such as PEI and CDP-imid can buffer the endocytic vesicles while PLL and CDP do not. Additionally, the PEGylated vectors reveal the same buffering capacity as their unstabilized variants. Here, the live cell, spatiotemporal mapping of these behaviors is demonstrated and, when combined with cell uptake and luciferase expression data, shows that there is not a correlation between buffering capacity and gene expression.
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| Additional Information: | © 2005 American Chemical Society. Received March 16, 2005; Revised Manuscript Received May 10, 2005. Publication Date (Web): June 28, 2005. We thank Dr. John Murphy for helpful discussions. R.P. Kulkarni acknowledges support from an NDSEG fellowship. We thank Insert Therapeutics, Inc., for partial support of this work. M. E. Davis is a consultant to and has financial interest in Insert Therapeutics, Inc. | |||||||||
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| Issue or Number: | 4 | |||||||||
| DOI: | 10.1021/bc050081u | |||||||||
| Record Number: | CaltechAUTHORS:20160226-114842424 | |||||||||
| Persistent URL: | https://resolver.caltech.edu/CaltechAUTHORS:20160226-114842424 | |||||||||
| Official Citation: | Single Cell Kinetics of Intracellular, Nonviral, Nucleic Acid Delivery Vehicle Acidification and Trafficking Rajan P. Kulkarni, Swaroop Mishra, Scott E. Fraser, and Mark E. Davis Bioconjugate Chemistry 2005 16 (4), 986-994 DOI: 10.1021/bc050081u | |||||||||
| Usage Policy: | No commercial reproduction, distribution, display or performance rights in this work are provided. | |||||||||
| ID Code: | 64803 | |||||||||
| Collection: | CaltechAUTHORS | |||||||||
| Deposited By: | Melissa Ray | |||||||||
| Deposited On: | 29 Feb 2016 18:55 | |||||||||
| Last Modified: | 10 Nov 2021 23:35 |
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