title: Two-photon fluorescence imaging super-enhanced by multishell nanophotonic particles, with application to subcellular pH.
abstract: 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.
authors.affiliation: BioPhysics, University of Michigan, 930 N. University Ave. Ann Arbor, MI 48109, USA.
authors.forename: Aniruddha
authors.initials: A
authors.lastname: Ray
authors.forename: Yong-Eun Koo
authors.initials: YE
authors.lastname: Lee
authors.forename: Gwangseong
authors.initials: G
authors.lastname: Kim
authors.forename: Raoul
authors.initials: R
authors.lastname: Kopelman
mesh_terms: D045744:Cell Line, Tumor; D005456:Fluorescent Dyes / Q000737:chemistry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007089:Image Enhancement / Q000379:methods; D036641:Microscopy, Fluorescence, Multiphoton / Q000379:methods*; D053758:Nanoparticles / Q000737:chemistry*; D061848:Optical Imaging / Q000379:methods*; D010316:Particle Size; D014961:X-Ray Diffraction
chemical_list: D005456:Fluorescent Dyes