Detection of low quantum yield fluorophores and improved imaging times using metallic nanoparticles

M. J. ROBERTI; LAURA C. ESTRADA; VALERIA LEVI; OSCAR E. MARTÍNEZ; PEDRO F. ARAMENDÍA

Mendoza

Congreso; 21st IAPS Conference; 2011

IAPS

Fluorophore-metal nanoparticle systems constitute excellent biosensors for applications in the field of cellular biology1. Gold NPs (AuNPs) can enhance the emission of fluorophores through interaction of the NP plasmon with the molecule absorption and emission moment2. In previous works3,4, we modeled this effect on AuNPs coupled to model fluorophores. We showed an enhancement in brightness and photostability for the fluorophores interacting with AuNPs, while the extent of this enhancement was spatially reduced up to 10 nm.Here, we address this enhancement effect in vitro and in cells using fluorescence microscopies. The enhancement in brightness in imaging experiments in vitro was assessed by using Au-SiO2 core-shell nanoparticles deposited on glass, and a ~13-fold enhanced contrast for weakly fluorescent molecules in the vicinity of the nanoparticles was observed. A significant increase in the fluorophore photostability rendering longer imaging times was obtained for fluorophores interacting with Au nanoparticles incorporated by endocytosis in cells. We employed AuNPs combined with different fluorophores, and easily internalized through endocytosis. We confirmed the enhancement of fluorescence on endocyted AuNPs interacting with fluorescently-labeled actin filaments, evidenced as an improved contrast and prolonged imaging times for those fluorophores associated to the filaments and at close vicinity to the nanoparticles4.This work shows the possibility of detecting intrinsic fluorescence of biomolecules and obtaining longer imaging times at the single molecule level, with subdiffraction resolution. The combination of AuNPs and fluorophores may be advantageously applied in multiple trafficking studies by stationary and time-solved microscopies, coupled to high resolution EM microscopy to characterize the localization and morphology of the targets.1. Shaoqin Liu and Zhiyong Tang. J. Mater. Chem. 20, 24–35 (2010).2. Ghosh, S. K. and T. Pal.. Phys. Chem. Chem. Phys. 11, 3831-3844 (2009).3. Estrada, L. C., P. F. Aramendía, and O. E. Martínez. Opt. Express 16, 20597-20602 (2008).4. L. Estrada, M. Julia Roberti, V. Levi, P. F. Aramendía, and O. Martínez. Manuscript in preparation.