Nanoscopía de fluorescencia tridimensional con resolución por debajo de 10 nm basada en la utilización de excitación súper-crítica

ALAN MARCELO SZALAI; BRUNO SIARRY; JERÓNIMO LUKIN; DAVID WILLIAMSON; NICOLAS UNSAIN; DAMIAN REFOJO; ALFREDO CÁCERES; MAURICIO PILO-PAIS; DYLAN OWEN; GUILLERMO ACUÑA; SABRINA SIMONCELLI; FERNANDO D. STEFANI

Buenos Aires

Encuentro; XIX Encuentro de superficies y materiales nanoestructurados.; 2019

Single-molecule localization microscopy enables far-field imaging with lateral resolution in the range of 10 to 20 nanometres, exploiting the fact that the centre position of a single-molecule's image can be determined with much higher accuracy than the size of that image itself. However, attaining the same level of resolution in the axial (third) dimension remains challenging. Here, we present Supercritical Illumination Microscopy Photometric z-Localization with Enhanced Resolution (SIMPLER), a photometric method to decode the axial position of single molecules in a total internal reflection fluorescence microscope. SIMPLER requires no hardware modification whatsoever to a conventional total internal reflection fluorescence microscope and complements any 2D single-molecule localization microscopy method to deliver 3D images with nearly isotropic nanometric resolution. Performance examples include SIMPLER-direct stochastic optical reconstruction microscopy images of the nuclear pore complex with sub-20 nm axial localization precision and visualization of microtubule cross-sections through SIMPLER-DNA points accumulation for imaging in nanoscale topography with sub-10 nm axial localization precision.