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Two‐Photon Fluorescence Imaging Super‐Enhanced by Multishell Nanophotonic Particles, with Application to Subcellular pH
Author(s) -
Ray Aniruddha,
Lee YongEun Koo,
Kim Gwangseong,
Kopelman Raoul
Publication year - 2012
Publication title -
small
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.785
H-Index - 236
eISSN - 1613-6829
pISSN - 1613-6810
DOI - 10.1002/smll.201102664
Subject(s) - photobleaching , fluorophore , fluorescence , two photon excitation microscopy , fluorescence in the life sciences , autofluorescence , materials science , microscopy , nanoprobe , fluorescence lifetime imaging microscopy , photochemistry , fluorescence microscope , nanoparticle , optoelectronics , nanotechnology , optics , chemistry , physics
A novel nanophotonic method for enhancing the two‐photon fluorescence signal of a fluorophore is presented. It utilizes the second harmonic (SH) of the exciting light generated by noble metal nanospheres in whose near‐field the dye molecules are placed, to further enhance the dye's fluorescence signal in addition to the usual metal‐enhanced fluorescence phenomenon. This method enables demonstration, for the first time, of two‐photon fluorescence enhancement inside a biological system, namely live cells. A multishell hydrogel nanoparticle containing a silver core, a protective citrate capping, which serves also as an excitation quenching inhibitor spacer, a pH indicator dye shell, and a polyacrylamide cladding are employed. Utilizing this technique, an enhancement of up to 20 times in the two‐photon fluorescence of the indicator dye is observed. Although a significant portion of the enhanced fluorescence signal is due to one‐photon processes accompanying the SH generation of the exciting light, this method preserves all the advantages of infrared‐excited, two‐photon microscopy: enhanced penetration depth, localized excitation, low photobleaching, low autofluorescence, and low cellular damage.