Subdiffraction Tracking of the Endocytic Pathway of Gold Nanoparticle-Fluorophore Systems

M. JULIA ROBERTI; L. ESTRADA; V. LEVI; O. MARTINEZ; P.F. ARAMENDIA

Salta, Argentina

Congreso; 3rd Latin American Protein Society Meeting; 2010

The Protein Society (http://www.proteinsociety.org)

Fluorophore-metal nanoparticle systems constitute excellent biosensors for applications in the field of cellular biology(1). Gold NPs (AuNPs) can enhance the emission of fluorophores through interaction of the NP plasmon with the molecule absorption and emission moment(2). In previous works(3,4), we modelled 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 effect in cells using fluorescence microscopies. 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 nanoparticles(4). We now aim to track endosomes with improved resolution using FCS microscopy. To this end, we have expressed a fluorescent quimera of the endosomal membrane protein LAMP-1, and exploit the enhancement of fluorescence upon AuNPs endocytosis. Our first results show correlation times that agree with those expected for LAMP-1 diffusing in the membrane.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. In preparation.