Ratiometric optical PEBBLE nanosensors for real-time magnesium ion concentrations inside viable cells.
This paper presents the development and characterization of a highly selective magnesium fluorescent optical nanosensor, made possible by PEBBLE (probe encapsulated by biologically localized embedding) technology. A ratiometric sensor has been developed by co-immobilizing a dye that is sensitive to and highly selective for magnesium, with a reference dye in a matrix. The sensors are prepared via a microemulsion polymerization process, which entraps the sensing components inside a polymer matrix. The resultant spherical sensors are approximately 40 nm in diameter. The Coumarin 343 (C343) dye, which by itself does not enter the cell, when immobilized in a PEBBLE is used as the magnesium-selective agent that provides the high and necessary selectivity over other intracellular ions, such as Ca2+, Na+, and K+. The dynamic range of these sensors was 1-30 mM, with a linear range from 1 to 10 mM, with a response time of <4 s. In contrast to free dye, these nano-optodes are not perturbed by proteins. They are fully reversible and exhibit minimal leaching and photobleaching over extended periods of time. In vitro intracellular changes in Mg2+ concentration were monitored in C6 glioma cells, which remained viable after PEBBLE delivery via gene gun injection. The selectivity for Mg2+ along with the biocompatibility of the matrix provides a new and reliable tool for intracellular magnesium measurements.
Department of Chemistry, The University of Michigan, Ann Arbor, Michigan 48109, USA.
Edwin J
EJ
Park
Murphy
M
Brasuel
Caleb
C
Behrend
Martin A
MA
Philbert
Raoul
R
Kopelman
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002412:Cations / Q000032:analysis; D045744:Cell Line, Tumor; D002470:Cell Survival; D003374:Coumarins; D005453:Fluorescence; D005910:Glioma / Q000737:chemistry / Q000378:metabolism / Q000473:pathology; D008274:Magnesium / Q000032:analysis*; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D055095:Optics and Photonics; D011108:Polymers; D051381:Rats; D013997:Time Factors
D002412:Cations; D003374:Coumarins; D011108:Polymers; D008274:Magnesium; D002118:Calcium