Three-dimensional total internal reflection fluorescence nanoscopy with sub-10 nm resolution

Here, we present a single-molecule localization microscopy (SMLM) analysis method that delivers sub-10 nm z-resolution when combined with 2D total internal reflection (TIR) fluorescence imaging via DNA point accumulation for imaging nanoscale topography (DNA-PAINT). Axial resolution is obtained from a precise measurement of the emission intensity of single molecules under evanescent field excitation. This method can be implemented on any conventional TIR wide-field microscope without modifications. We validate this approach by resolving the periodicity of alpha-tubulin assembly in microtubules, demonstrating isotropic resolution below 8 nm.

[1]  Ambrose Ej A surface contact microscope for the study of cell movements. , 1956 .

[2]  E. J. Ambrose The movements of fibrocytes. , 1961, Experimental cell research.

[3]  W. Lukosz,et al.  Light emission by magnetic and electric dipoles close to a plane dielectric interface. II. Radiation patterns of perpendicular oriented dipoles , 1977 .

[4]  W. Lukosz,et al.  Light emission by magnetic and electric dipoles close to a plane interface. I. Total radiated power , 1977 .

[5]  W. Lukosz Light emission by magnetic and electric dipoles close to a plane dielectric interface. III. Radiation patterns of dipoles with arbitrary orientation , 1979 .

[6]  D. Axelrod Cell-substrate contacts illuminated by total internal reflection fluorescence , 1981, The Journal of cell biology.

[7]  D. Taylor,et al.  Structural organization of interphase 3T3 fibroblasts studied by total internal reflection fluorescence microscopy , 1985, The Journal of cell biology.

[8]  D. Axelrod,et al.  Fluorescence emission at dielectric and metal-film interfaces , 1987 .

[9]  D. Axelrod,et al.  Emission of fluorescence at an interface. , 1989, Methods in cell biology.

[10]  S. Hell,et al.  Properties of a 4Pi confocal fluorescence microscope , 1992 .

[11]  G. Truskey,et al.  Quantitative analysis of variable‐angle total internal reflection fluorescence microscopy (VA‐TIRFM) of cell/substrate contacts , 1994, Journal of microscopy.

[12]  G. Omann,et al.  Membrane-proximal calcium transients in stimulated neutrophils detected by total internal reflection fluorescence. , 1996, Biophysical journal.

[13]  B. Ölveczky,et al.  Mapping fluorophore distributions in three dimensions by quantitative multiple angle-total internal reflection fluorescence microscopy. , 1997, Biophysical journal.

[14]  D. Loerke,et al.  The last few milliseconds in the life of a secretory granule , 1998, European Biophysics Journal.

[15]  W. Almers,et al.  Tracking single secretory granules in live chromaffin cells by evanescent-field fluorescence microscopy. , 1999, Biophysical journal.

[16]  C. Martínez-A,et al.  Distribution and characteristics of betaII tubulin-enriched microtubules in interphase cells. , 1999, Experimental cell research.

[17]  A. Rohrbach,et al.  Observing secretory granules with a multiangle evanescent wave microscope. , 2000, Biophysical journal.

[18]  D. Loerke,et al.  Quantifying axial secretory-granule motion with variable-angle evanescent-field excitation , 2002, Journal of Neuroscience Methods.

[19]  Julio M Fernandez,et al.  Simultaneous atomic force microscope and fluorescence measurements of protein unfolding using a calibrated evanescent wave. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[20]  Michael D. Mason,et al.  Ultra-high resolution imaging by fluorescence photoactivation localization microscopy. , 2006, Biophysical journal.

[21]  Michael J Rust,et al.  Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM) , 2006, Nature Methods.

[22]  D. Axelrod,et al.  Direct measurement of the evanescent field profile produced by objective-based total internal reflection fluorescence. , 2006, Journal of biomedical optics.

[23]  R. Hochstrasser,et al.  Wide-field subdiffraction imaging by accumulated binding of diffusing probes , 2006, Proceedings of the National Academy of Sciences.

[24]  H. Ewers,et al.  Even illumination in total internal reflection fluorescence microscopy using laser light , 2008, Microscopy research and technique.

[25]  S. Hell,et al.  Spherical nanosized focal spot unravels the interior of cells , 2008, Nature Methods.

[26]  Mark Bates,et al.  Three-Dimensional Super-Resolution Imaging by Stochastic Optical Reconstruction Microscopy , 2008, Science.

[27]  T. Kirchhausen,et al.  Differential evanescence nanometry: live-cell fluorescence measurements with 10-nm axial resolution on the plasma membrane. , 2008, Biophysical journal.

[28]  M. Heilemann,et al.  Subdiffraction-resolution fluorescence imaging with conventional fluorescent probes. , 2008, Angewandte Chemie.

[29]  S. Diez,et al.  TIRF microscopy evanescent field calibration using tilted fluorescent microtubules , 2009, Journal of microscopy.

[30]  J. Lippincott-Schwartz,et al.  Interferometric fluorescent super-resolution microscopy resolves 3D cellular ultrastructure , 2009, Proceedings of the National Academy of Sciences.

[31]  F. Simmel,et al.  Single-molecule kinetics and super-resolution microscopy by fluorescence imaging of transient binding on DNA origami. , 2010, Nano letters.

[32]  Wei Sun,et al.  Autocalibrated scanning-angle prism-type total internal reflection fluorescence microscopy for nanometer-precision axial position determination. , 2010, Analytical chemistry.

[33]  James S. Duncan,et al.  3-D Reconstruction of Microtubules From Multi-Angle Total Internal Reflection Fluorescence Microscopy Using Bayesian Framework , 2011, IEEE Transactions on Image Processing.

[34]  S. Hell,et al.  Two-color nanoscopy of three-dimensional volumes by 4Pi detection of stochastically switched fluorophores , 2011, Nature Methods.

[35]  Guy M. Hagen,et al.  ThunderSTORM: a comprehensive ImageJ plug-in for PALM and STORM data analysis and super-resolution imaging , 2014, Bioinform..

[36]  J. Boulanger,et al.  Fast high-resolution 3D total internal reflection fluorescence microscopy by incidence angle scanning and azimuthal averaging , 2014, Proceedings of the National Academy of Sciences.

[37]  J. Enderlein,et al.  Metal-induced energy transfer for live cell nanoscopy , 2014, Nature Photonics.

[38]  E. Fort,et al.  Direct optical nanoscopy with axially localized detection , 2014, Nature Photonics.

[39]  Simon J. Herr,et al.  isoSTED nanoscopy with intrinsic beam alignment. , 2015, Optics express.

[40]  Federico M Barabas,et al.  Note: Tormenta: An open source Python-powered control software for camera based optical microscopy. , 2016, The Review of scientific instruments.

[41]  G. Patterson,et al.  Axial superresolution via multiangle TIRF microscopy with sequential imaging and photobleaching , 2016, Proceedings of the National Academy of Sciences.

[42]  Henrik Flyvbjerg,et al.  “Calibration-on-the-spot”: How to calibrate an EMCCD camera from its images , 2016, Scientific Reports.

[43]  R. Alon,et al.  Three-dimensional localization of T-cell receptors in relation to microvilli using a combination of superresolution microscopies , 2016, Proceedings of the National Academy of Sciences.

[44]  Jordan R. Myers,et al.  Ultra-High Resolution 3D Imaging of Whole Cells , 2016, Cell.

[45]  J. Elf,et al.  Nanometer resolution imaging and tracking of fluorescent molecules with minimal photon fluxes , 2016, Science.

[46]  M. Sauer,et al.  Photometry unlocks 3D information from 2D localization microscopy data , 2016, Nature Methods.

[47]  S. Hell,et al.  Fluorescence nanoscopy in cell biology , 2017, Nature Reviews Molecular Cell Biology.

[48]  Maximilian T. Strauss,et al.  Super-resolution microscopy with DNA-PAINT , 2017, Nature Protocols.

[49]  A. Lambacher,et al.  Direct characterization of the evanescent field in objective-type total internal reflection fluorescence microscopy. , 2018, Optics express.

[50]  J. Enderlein,et al.  Three-dimensional single-molecule localization with nanometer accuracy using Metal-Induced Energy Transfer (MIET) imaging. , 2018, The Journal of chemical physics.

[51]  M. Oheim,et al.  Calibrating Evanescent-Wave Penetration Depths for Biological TIRF Microscopy. , 2019, Biophysical journal.

[52]  Vilma Jimenez Sabinina,et al.  Optimal 3D single-molecule localization in real time using experimental point spread functions , 2018, Nature Methods.

[53]  P. Tinnefeld,et al.  Distance Dependence of Single-Molecule Energy Transfer to Graphene Measured with DNA Origami Nanopositioners. , 2019, Nano letters.

[54]  J. Enderlein,et al.  Graphene-based metal-induced energy transfer for sub-nanometre optical localization , 2019, Nature Photonics.

[55]  Maximilian T. Strauss,et al.  Direct Visualization of Single Nuclear Pore Complex Proteins Using Genetically‐Encoded Probes for DNA‐PAINT , 2019, Angewandte Chemie.

[56]  Ulf Matti,et al.  Nuclear pores as versatile reference standards for quantitative superresolution microscopy , 2019, Nature Methods.