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Single Cell Kinetics of Intracellular, Nonviral, Nucleic Acid Delivery Vehicle Acidification and Trafficking

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.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1021/bc050081uDOIArticle
http://pubs.acs.org/doi/abs/10.1021/bc050081uPublisherArticle
ORCID:
AuthorORCID
Mishra, Swaroop0000-0002-2665-5385
Fraser, Scott E.0000-0002-5377-0223
Davis, Mark E.0000-0001-8294-1477
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.
Funders:
Funding AgencyGrant Number
National Defense Science and Engineering Graduate (NDSEG) FellowshipUNSPECIFIED
Insert Therapeutics, Inc.UNSPECIFIED
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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