Suchergebnisse

In the replacement of genetic probes, there is increasing interest in labeling living cells with high-quality extrinsic labels, which avoid over-expression artifacts and are available in a wide spectral range. This calls for a broadly applicable technology that can deliver such labels unambiguously to the cytosol of living cells. Here, we demonstrate that nanoparticle-sensitized photoporation can be used to this end as an emerging intracellular delivery technique. We replace the traditionally used gold nanoparticles with graphene nanoparticles as photothermal sensitizers to permeabilize the cell membrane upon laser irradiation. We demonstrate that the enhanced thermal stability of graphene quantum dots allows the formation of multiple vapor nanobubbles upon irradiation with short laser pulses, allowing the delivery of a variety of extrinsic cell labels efficiently and homogeneously into live cells. We demonstrate high-quality time-lapse imaging with confocal, total internal reflection fluorescence (TIRF), and Airyscan super-resolution microscopy. As the entire procedure is readily compatible with fluorescence (super resolution) microscopy, photoporation with graphene quantum dots has the potential to become the long-awaited generic platform for controlled intracellular delivery of fluorescent labels for live-cell imaging. ; K.B. acknowledges financial support from the European Research Council (ERC) under the European Union's Horizon 2020 research and innovation program (grant agreement No 648124) and from the Ghent University Special Research Fund (01B04912) with gratitude. J.L. gratefully acknowledges the financial support from the China Scholarship Council (CSC) (201506750012) and the Special Research Fund from Ghent University (01SC1416). R.X. gratefully acknowledges the financial support from the China Scholarship Council (CSC) (2010634103). H.B. acknowledges funding from the Research Foundation Flanders (Fonds Wetenschappelijk Onderzoek, FWO) for a doctoral fellowship (11ZB115N). E.T. acknowledges funding from the Agency for Innovation by Science and Technology (IWT). S.S. and R.B. acknowledge financial support from the Centre National de la Recherche Scientifique (CNRS), the University of Lille, the Hauts-de-France region, the CPER "Photonics for Society", and the EU union through FLAG-ERA JTC 2015-Graphtivity and the Marie Sklodowska-Curie action (H2020-MSCA-RISE-2015, PANG-690836). M.A. acknowledges the support by the FWO Research Community "Scanning and Wide Field Microscopy of (Bio)-organic Systems" and the Province of Limburg (Belgium) for the financial support within the tUL IMPULS FASE II program, allowing for the upgrade of the laser source used in this work. We would thank the VIB BioImaging Core for the use of the microscopy equipment.