Near field fluorescence fluctuation spectroscopy with gold nanoparticles

LAURA ESTRADA; PEDRO F. ARAMENDÍA; OSCAR E. MARTÍNEZ

La Serena, Chile

Congreso; X Encuentro Latinoamerican o de Fotoquímica y Fotobiología; 2010

The interaction of chromophores with metal nanoparticles (MNPs) has been extensively studied in recent years. Fluorophores in the close vicinity of metal nanoparticles are interesting systems to develop sensors, especially in the fields of cellular tracking and imaging. Fluorescence enhancement of a fluorophore in the nanometer distance scale from a metal nanoparticle is well documented, a phenomenon attributed to the near field interaction of the molecule oscillating transition dipole with a characteristic resonance frequency of the free electrons of the nanostructure. MNPs can modify the emission of fluorescent molecules by increasing the excitation and emission rates (plasmonic enhancement), as well as by modifying the non‑radiative energy transfer from the molecule to the particle (quenching effect). These processes are influenced by the MNP size, shape, and material; the fluorophore absorption and emission properties, quantum yield, and orientation; MNP‑fluorophore distance; and the dielectric constant of the surrounding medium. The modification in excitation, emission, and non‑radiative energy transfer rates is distance dependent and, as a result, an overall fluorescence quenching results at very short distance from the MNP, an emission enhancement is observed at intermediate distances, and no influence results, for a molecule placed at a distance greater than about twice the MNP radius. It is clear that a careful analysis of the effects of the actual MNP‑fluorophore characteristics on the fluorescence emission is of broad interest to find conditions that optimize a desirable property of the probe. For a rational design of a MNP‑fluorophore hybrid system it is necessary to take into account all the variables that modify the chromophore photophysics and that can lead to the desired fluorescence enhancement. We summarize a careful study on the influence of all the mentioned variables on the photophysics of molecules in the close vicinity to MNPs and we present results of three experiments using the emission enhancement by 40 nm radius AuNP irradiated in the green in a confocal microscope. 1) Fluorescence fluctuation spectroscopy that shows a reduction of a factor of 104 in the observation volume. 2) Localization experiments that show the detectability of Rose Bengal with a great contrast. 3) Tracking experiments in cells that show an increase in tracking time and persistence against photobleaching.