Three-dimensional total-internal reflection fluorescence nanoscopy with nanometric axial resolution by photometric localization of single molecules

Single-molecule localization microscopy (SMLM) 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 SIMPLER, a photometric method to decode the axial position of single molecules in a total internal reflection fluorescence (TIRF) microscope. SIMPLER requires no hardware modification whatsoever to a conventional TIRF microscope, and complements any 2D SMLM method to deliver 3D images with nearly isotropic nanometric resolution. Examples of the performance of SIMPLER include the visualization of the nuclear pore complex through dSTORM with sub-20 nm resolution and of microtubule cross-sections resolved with sub-10 nm through DNA-PAINT.

[1]  E. J. Ambrose.  A Surface Contact Microscope for the study of Cell Movements , 1956, Nature.

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

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

[4]  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 .

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

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

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

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

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

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

[11]  G. Truskey,et al.  Total internal reflection fluorescence (TIRF) microscopy. I. Modelling cell contact region fluorescence. , 1990, Journal of cell science.

[12]  G. Truskey,et al.  Total internal reflection fluorescence microscopy (TIRFM). II. Topographical mapping of relative cell/substratum separation distances. , 1992, Journal of cell science.

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

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

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

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

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

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

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

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

[21]  W. Almers,et al.  Imaging actin and dynamin recruitment during invagination of single clathrin-coated pits , 2002, Nature Cell Biology.

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

[23]  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.

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

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

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

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

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

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

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

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

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

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

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

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

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

[37]  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.

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

[39]  H. Ewers,et al.  A simple, versatile method for GFP-based super-resolution microscopy via nanobodies , 2012, Nature Methods.

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

[41]  M. Mund,et al.  3D superresolution microscopy by supercritical angle detection , 2014 .

[42]  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.

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

[44]  M. Mund,et al.  3D superresolution microscopy by supercritical angle detection. , 2014, Optics express.

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

[46]  R. Vale,et al.  Total Internal Reflection Fluorescence Microscopy. , 2015, Cold Spring Harbor protocols.

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

[48]  S. Hell.  Nanoscopy with Focused Light (Nobel Lecture). , 2015, Angewandte Chemie.

[49]  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.

[50]  N. Hawkes,et al.  In situ wavelength calibration of the edge CXS spectrometers on JET. , 2016, The Review of scientific instruments.

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

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

[53]  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.

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

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

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

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

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

[59]  Ulf Matti,et al.  Optimal 3D single-molecule localization in real time using experimental point spread functions , 2018, Nature Methods.

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

[61]  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.

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

[63]  A. Descloux,et al.  Parameter-free image resolution estimation based on decorrelation analysis , 2019, Nature Methods.

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

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

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

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

[68]  J. Ellenberg,et al.  MINFLUX nanoscopy delivers 3D multicolor nanometer resolution in cells , 2020, Nature Methods.

[69]  Three-dimensional total-internal reflection fluorescence nanoscopy with nanometric axial resolution by photometric localization of single molecules , 2021, Nature communications.