MAR 17, 2020 7:00 AM PDT

Time-Resolved STED Microscopy

Sponsored by: Leica Microsystems
C.E. Credits: P.A.C.E. CE Florida CE
Speaker
  • Senior Research Tenured Molecular Microscopy and Spectroscopy, Center for Human Technology, Istituto Italiano di Tecnologia, Genoa, Italy
    Biography
      I studied computer science at the University of Genoa and received my Diploma cum laude in 2003 (advised by Prof. Mario Bertero and Prof. Patrizia Boccacci). From 2003 to 2007 I worked at the Laboratory of Advance Microscopy and Spectroscopy where I received my Ph.D. (advised by Prof. Alberto Diaspro) about image processing and analysis for fluorescence microscopy.
      From 2008 to 2011 I worked as a post-doctoral fellow at the Department of NanoBiophotonics (headed by Prof. Stefan W. Hell, Nobel Laureate in 2014), MPI for Biophysical Chemistry; where I developed a new approach, based on the temporal analysis of the fluorescence signal, which allows stimulated emission deletion (STED) microscopy to achieve tens of nanometres spatial resolution with a substantial reduction of the dose of light requested, thus opening the effective application of STED microscopy with fluorescent proteins and living cells. Currently, all commercially available STED microscopes implement this method - called gated- STED. This work pioneered the use of the temporal information channel of a microscope, such as the ability of measuring the fluorescence signal dynamics at the nanoseconds scale, for improving the microscope spatial-resolution.
      Since May 2011 I work in the Department of NanoPhysics (headed by Prof. Alberto Diaspro), Italian Institute of Technology, where in September 2013 I obtained a Researcher position, in May 2016 I became principal investigator of the Molecular Microscopy and Spectroscopy research line, and in December 2019 I was granted tenure. I am also Adjunct Professor at the University of Genoa. While constantly working on STED microscopy, and on its combination with fluorescence-correlation spectroscopy (STED-FCS), I started also developing novel single-photon-avalanche diode (SPAD) array detectors for fluorescence microscopy. This research has recently evolved in the invention of a new scanning microscopy technique able to double the spatial resolution of conventional microscopy, while maintain live-cell, multi-color, and three-dimension imaging capabilities and adding time-resolved spectroscopy/imaging (e.g., fluorescence lifetime, intensity fluctuation analysis, anti-bunching). This technique has introduced a new paradigm in fluorescence microscopy which proposes to unlock the secret carried by each single-photon, thus moving from the era of single-molecule microscopy to the era of single-photon microscopy.
      My work has been supported by grants from various agencies such as the Fondazione San Paolo (Italy), the Marie Sklodowska Curie Actions (MSCA) and the European Research Council (ERC).
      I am co-founder and scientific advisor of the Genoa Instruments spin-off company. The spin-off is dedicated to launch a software and hardware add-on system which will transform any conventional confocal microscope into a high-resolution single-photon microscope.

    Abstract
    DATE:  March 17, 2020
    TIME:  7:00am PT, 10:00am ET
     
    Introduced more than 30 years ago, stimulated emission depletion (STED) microscopy has raised to a standard and widely used method for imaging in the life sciences. Thanks to continuous technological progress, STED microscopy can now provide effective sub-diffraction spatial resolution, while preserving most of the useful aspects of fluorescence microscopy, such as optical sectioning, molecular specificity and sensitivity, and live-cell compatibility.
     
    This seminar starts by presenting the basic principles of STED microscopy and the three significant conditions required for its implementation, specifically the spatial, spectral, and temporal conditions. Towards these conditions, the seminar introduces the initial limitations and problems of STED microscopy and how modern STED microscopy architectures have circumvented these limitations. In particular, the webinar shows how the high illumination intensity - required to achieve tens of nanometer spatial resolution - can be mitigated by time-resolved STED microscopy, namely by recording and analyzing the temporal dynamics of the fluorescence signal (i.e., the fluorescence lifetime). In this context, the seminar explains and compares the time-gated and, more recently, separation-of-photons-by-lifetime-tuning (SPLIT) STED microscopy implementations.
     
    Learning Objectives:  
    • Spatial, Spectral and Temporal Conditions for STED microscopy
    • Mitigating high illumination intensity with time-resolved STED microscopy taking advantage of fluorescence lifetime analysis (FLIM)
    • Comparison of time-gated and Separation-of-Photons-by-Lifetime (SPLIT) STED applications

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