Photonics West 2022 in San Francisco
Starts Saturday
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We are happy to announce three invited lectures. Michelle Digman, University of California Irvine, and Falk Schneider, University of Southern California (Scott Fraser group), will highlight advanced biological applications and latest results using various methods of fluorescence (raster) correlation spectroscopy. Peter Dahlberg from Stanford will present exciting results correlating cryogenic Super-Resolution Fluorescence with Cryogenic Electron Tomography.


In the focus of this conference are all fields of optical single molecule spectroscopy and super resolution imaging, ranging from fundamental physics, technical and methodological questions, towards applications in chemical, biological and biomedical research as well as medical diagnostics. It provides a state-of-the-art interdisciplinary forum for information exchange on new technological developments, advanced applications, and fundamental questions of the field.

Ultra-sensitive spectroscopic techniques have become an important tool in fundamental biological and biomedical research, allowing study of the function and interaction of individual biomolecules. Improving and extending the existing arsenal of techniques for studying specific biophysical and biochemical questions on a single molecule level is of paramount interest for the life-science community.

This conference puts special emphasis on time resolved methods of fluorescence spectroscopy and imaging which allow for investigating not only structural properties but also the function of molecular processes, down to the single molecule level. Therefore, we encourage to submit work related also to Fluorescence Lifetime Imaging (FLIM), Advanced single-molecule techniques such as Fluorescence Correlation Spectroscopy (FCS), Fluorescence Coincidence Analysis or single-molecule burst analysis are also favorite subjects of this conference. In particular Förster resonance energy transfer (FRET) analysis frequently benefits from theses time-resolved methods and this conference will be an excellent platform to discuss their application at the molecular level.

A topic of particular interest has become the employment of the single-molecule nature of fluorescence excitation and emission to achieve sub-diffraction superresolution in fluorescence microscopy. It has opened previously unknown opportunities to image live cells in the optical far field with unprecedented optical resolution. This resulted in new microscopy modalities such as Stimulated Emission Depletion (STED) microscopy, single molecule localization microscopy (PALM, STORM, dSTORM, GSD-IM), stochastic optical fluctuation microscopy (SOFI), or structured illumination microscopy (SIM) and imaging scanning microscopy (ISM) techniques. The conference provides an interdisciplinary platform for these new and exciting developments in fluorescence imaging.

The need for ultrasensitive and specific biomedical diagnostics requires development of optical and photonic detection/sensing technologies capable of reaching the single molecule level. The technical challenges to rapidly and specifically detect chemical and biological agents at minimal concentration levels are enormous and largely yet to be realized. All spectroscopic techniques (optical spectroscopy, fluorescence spectroscopy, elastic scattering, Raman scattering, IR spectroscopy, terahertz spectroscopy) as well as the chemical and biological sciences themselves including genetically encoded fluorescent markers and (photoswitchable) labels, are potentially critical components for a multidisciplinary approach to ultrasensitive sensing and diagnostics.

Invited and contributed papers are solicited concerning, but not limited to, the following areas:
PICOQUANT YOUNG INVESTIGATOR AWARD
Young scientists (age 30 or below and not yet full faculty members) are encouraged to participate in this best paper competition, which offers a $750 USD cash award. Participants must be both the primary author and presenter of an accepted abstract to be eligible. Please select "PicoQuant Young Investigator Award" as the last Topic in the abstract submission wizard in order to be considered. This award is sponsored by PicoQuant GmbH Berlin and presented Sunday afternoon.
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In progress – view active session
Conference 11967

Single Molecule Spectroscopy and Superresolution Imaging XV

22 - 23 January 2022 | Room 201 (Level 2 South)
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  • Welcome and Introduction
  • 1: New Techniques in Superresolution Microscopy
  • 2: FLIM, FRET & FCS/Correlation
  • 3: New Technologies & Biological Applications
  • 4: Superresolution Microscopy/Nanoscopy I
  • 5: Multimodal and Correlative Technologies
  • 6: Superresolution Microscopy/Nanoscopy II
  • 7: Young Investigator Award Presentation
  • Posters-Sunday
Information

Check the conference schedule frequently for updates | Presentation times are subject to change

  • Presenters: Please inform SPIE of any changes by 7 January
  • Presentation times will be finalized on 19 January
Welcome and Introduction
22 January 2022 • 10:10 AM - 10:15 AM PST | Room 201 (Level 2 South)
Welcome remarks and introduction by conference chair Rainer Erdmann.
Session 1: New Techniques in Superresolution Microscopy
22 January 2022 • 10:15 AM - 10:35 AM PST | Room 201 (Level 2 South)
Session Chair: Rainer Erdmann, PicoQuant GmbH (Germany)
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Author(s): Shih-Huan Huang, National Taiwan Univ. (Taiwan); Kohei Otomo, National Institute for Physiological Sciences, National Institutes of Natural Sciences (Japan), Exploratory Research Ctr. on Life and Living Systems, National Institutes of Natural Sciences (Japan), School of Life Science, The Graduate Univ. for Advanced Studies (Japan); Shiu-Feng Cheng, National Taiwan Univ. (Taiwan); Kuo-Chuan Chao, Brain Research Ctr., National Tsing Hua Univ. (Taiwan); Yan-Wei Chen, Shun-Chi Wu, National Tsing Hua Univ. (Taiwan); Li-An Chu, Brain Research Ctr., National Tsing Hua Univ. (Taiwan); Ann-Shyn Chiang, Brain Research Ctr., National Tsing Hua Univ. (Taiwan), Kavli Institute for Brain and Mind, Univ. of California, San Diego (United States), Institute of Systems Neuroscience, National Tsing Hua Univ. (Taiwan); Tomomi Nemoto, National Institute for Physiological Sciences, National Institutes of Natural Sciences (Japan), Exploratory Research Ctr. on Life and Living Systems, National Institutes of Natural Sciences (Japan), School of Life Science, The Graduate Univ. for Advanced Studies (Japan); Shi-Wei Chu, Brain Research Ctr., National Tsing Hua Univ. (Taiwan), Molecular Imaging Ctr., National Taiwan Univ. (Taiwan)
22 January 2022 • 10:15 AM - 10:35 AM PST | Room 201 (Level 2 South)
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Recently, whole-Drosophila-brain structural mapping has been demonstrated with ~50-nm resolution, but not in vivo. In this work, we combine two-photon excitation into a spinning disk confocal setup to enable ~500 μm imaging depths in uncleared mouse brains (~100 μm in Drosophila), and wide-field optical sectioned detection, respectively. Through integration of photo-activated localization microscopy (PALM), which also reduces photodamage of the tissue, we achieve resolution down to ~50 nm in an intact Drosophila brain, pushing to the limit of imaging depth and resolution. Our work paves the way toward in vivo functional connectome studies.
Session 2: FLIM, FRET & FCS/Correlation
22 January 2022 • 10:35 AM - 12:15 PM PST | Room 201 (Level 2 South)
Session Chair: Linnea Olofsson, PicoQuant Photonics North America, Inc. (United States)
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Author(s): Michelle Digman, Univ. of California, Irvine (United States)
22 January 2022 • 10:35 AM - 11:05 AM PST | Room 201 (Level 2 South)
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The hallmark of metabolic alteration of increase glycolysis, i.e. Warburg effect, in cancer cells together with atypical extracellular matrix structure may be responsible for tumor cell aggressiveness and drug resistance. Here we apply the phasor approach technique in fluorescence lifetime imaging microscopy (FLIM) as a novel method to measure metabolic alteration as a function of ECM mechanics. We imaged and compared triple-negative breast cancer (TNBC) cells to non-cancerous cells on various ECM stiffness. Dysregulation of mitochondrial motion may contribute to the fueling of bioenergy demands in metastatic cancer. To measure mitochondria motion and analyze their fusion and fission events, we developed a new algorithm called “mitometer” that is unbiased, and allows for automated segmentation and tracking of mitochondria in live cell 2D and 3D time-lapse images.
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Author(s): Falk Schneider, The Univ. of Southern California (United States); Pablo F. Cespedes, Univ. of Oxford (United Kingdom); Erdinc Sezgin, Karolinska Institute (Sweden); Marco Fritzsche, Univ. of Oxford (United Kingdom); Scott E. Fraser, The Univ. of Southern California (United States)
22 January 2022 • 11:05 AM - 11:35 AM PST | Room 201 (Level 2 South)
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Fluorescence fluctuation spectroscopy (FFS) offers a window to observe and investigate dynamics of bio-molecular organisation. Furthermore, FFS acquisitions uniquely allow obtaining insights into other properties of the sample such as concentration and brightness (counts per molecule) of the fluorescently labelled species simultaneously. Here, we exploit the large number of measurements and statistics obtained from scanning fluorescence fluctuation spectroscopy (sFFS) acquisitions to elucidate bio-molecular organisation. We demonstrate the versatility and sensitivity of the approach using computer simulations, synthetic biological membranes and living cells.
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Author(s): Ingo Gregor, Arindam Ghosh, Tao Chen, Georg-August-Univ. Göttingen (Germany); Sufi O. Raja, Duke Soft Matter Ctr., Duke Univ. (United States); Alexey I. Chizhik, Georg-August-Univ. Göttingen (Germany); Christoph F. Schmidt, Duke Soft Matter Ctr., Duke Univ. (United States); Jörg Enderlein, Georg-August-Univ. Göttingen (Germany)
22 January 2022 • 11:35 AM - 11:55 AM PST | Room 201 (Level 2 South)
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Graphene-induced energy transfer (GIET) was recently introduced for sub-nanometric axial localization of fluorescent molecules. GIET exploits the near-field energy transfer from an excited fluorophore to a single sheet of graphene. This alters the fluorescence decay-time of the emitter and can be easily determined by fluorescence lifetime imaging microscopy (FLIM). The axial resolution of GIET implies to study of biological membranes. We present the measurement of the thickness of synthetic model membranes and demonstrate changes upon the addition of cholesterol. Furthermore, we are able to show the flipping of lipids from one leaflet to the other and determine the rates of this dynamics. In addition, we used GIET for mapping quasi-stationary states of the mitochondrial membranes before and during ATP synthesis. Upon activation, the inner membrane clearly approaches the outer membrane and the inter-membrane space is reduced by ∼2 nm.
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Author(s): Donald Ferschweiler, Rio Hondo Community College (United States), Univ. of California, Los Angeles (United States); Maya Segal, Xavier Michalet, Shimon Weiss, Univ. of California, Los Angeles (United States)
22 January 2022 • 11:55 AM - 12:15 PM PST | Room 201 (Level 2 South)
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Photon-HDF5 is an open-source and open file format for storing photon-counting data from single molecule microscopy experiments, introduced to simplify data exchange and increase the reproducibility of data analysis. Part of the Photon-HDF5 ecosystem, is phconvert, an extensible python library that allows converting proprietary formats into Photon-HDF5 files. However, its use requires some proficiency with command line instructions, the python programming language, and the YAML markup format. This creates a significant barrier for potential users without that expertise, but who want to benefit from the advantages of releasing their files in an open format. In this work, we present a GUI that lowers this barrier, thus simplifying the use of Photon-HDF5. This tool uses the phconvert python library to convert data files originally saved in proprietary data formats to Photon-HDF5 files, without users having to write a single line of code. Because reproducible analyses depend on essential experimental information, such as laser power or sample description, the GUI also includes (currently limited) functionality to associate valid metadata with the converted file, without having to write any YAML. Finally, the GUI includes several productivity-enhancing features such as whole-directory batch conversion and the ability to re-run a failed batch, only converting the files that could not be converted in the previous run.
Break
Lunch Break 12:15 PM - 1:35 PM
Session 3: New Technologies & Biological Applications
22 January 2022 • 1:35 PM - 2:55 PM PST | Room 201 (Level 2 South)
Session Chair: Rainer Erdmann, PicoQuant GmbH (Germany)
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Author(s): Serena Farina, Ivan Labanca, Giulia Acconcia, Politecnico di Milano (Italy); Alberto Ghezzi, Politecnico di Milano (Italy), Consiglio Nazionale delle Ricerche (Italy); Andrea Farina, Consiglio Nazionale delle Ricerche (Italy); Cosimo D'Andrea, Politecnico di Milano (Italy), Center for Nano Science and Technology at PoliMi (Italy); Ivan Rech, Politecnico di Milano (Italy)
22 January 2022 • 1:35 PM - 1:55 PM PST | Room 201 (Level 2 South)
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Time-Correlated Single-Photon Counting (TCSPC) is a time-resolved and ultrasensitive technique, providing the analysis of optical pulses in different domains. Unfortunately, its speed is strongly limited by pile-up distortion phenomena happening at high photon rates. Previously, we demonstrated that a perfect matching between detector dead time and laser period allows us to overcome this limitation, still maintaining negligible distortion. In this work, we present the design, characterization and experimental validation of a single-channel TCSPC system implementing the proposed idea. We carried out on-field fluorescence measurements employing the newly developed system, achieving high acquisition speed (32 Mcps) with extremely low lifetime distortion.
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Author(s): Luis Morales, Max-Planck-Institut für die Physik des Lichts (Germany); Julian Folz, Claus Seidel, Heinrich-Heine-Univ. Düsseldorf (Germany); Stephan J. Götzinger, Vahid Sandoghdar, Max-Planck-Institut für die Physik des Lichts (Germany); Ralf Kühnemuth, Suren Felekyan, Heinrich-Heine-Univ. Düsseldorf (Germany)
22 January 2022 • 1:55 PM - 2:15 PM PST | Room 201 (Level 2 South)
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In this work, we developed a planar dielectric antenna for analytes diffusing in aqueous solution. The so-called optofluidic antenna can collect more than 86% of all photons from a randomly oriented dipole-like emitter. The antenna involves a sub-micrometer water channel capped with air where the analytes are interrogated. The small dimension of the water channel in combination with the water/air interface confines the motion of the analytes, resulting in a slowing down of the translational diffusion. We characterize the photonic properties of the optofluidic antenna by investigating different dye molecules using fluorescence correlation spectroscopy. Moreover, we demonstrate the performance of our antenna by studying the dynamical behavior of the Holliday junction (HJ) at the single-molecule level using multiparameter fluorescence detection, which allows us to identify the HJ’s different FRET states in real-time.
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Author(s): Johanna Rahm, Sebastian Malkusch, Goethe-Univ. Frankfurt am Main (Germany); Ulrike Endesfelder, Rheinische Friedrich-Wilhelms-Univ. Bonn (Germany); Marina S. Dietz, Mike Heilemann, Goethe-Univ. Frankfurt am Main (Germany)
22 January 2022 • 2:15 PM - 2:35 PM PST | Room 201 (Level 2 South)
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We report segmentation analysis for single-particle tracking data of membrane receptors in living cells. Using a mean-squared displacement analysis, we classify trajectory segments into immobile, confined diffusing, and freely diffusing states, and extract the occurrence of transitions between these states in single trajectories. Using this method, we analyzed the diffusion of membrane receptor MET in untreated and ligand-treated living cells. We find that ligand treatment increased the frequency of transitions into the immobilie state. We also find that the confined diffusion state acts as an intermediate between immobile and free diffusion, in line with the process of receptor internalization.
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Author(s): Pavel N. Melentiev, Institute of Spectroscopy (Russian Federation); Rinat O. Esenaliev, The Univ. of Texas Medical Branch (United States); Lina V. Son, Denis S. Kudryavtsev, Igor E. Kasheverov, Victor I. Tsetlin, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry (Russian Federation); Victor I. Balykin, Institute of Spectroscopy (Russian Federation)
22 January 2022 • 2:35 PM - 2:55 PM PST | Room 201 (Level 2 South)
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Detection and imaging of single molecules have always been important tasks in fundamental science as well as practical applications. In this work we developed a novel approach to ultra-sensitive, ultra-fast detection and imaging of single molecules. Our approach allowed for detection of single troponin-T (cTnT) molecules (a clinically important marker for cardiac muscle damage) in human blood serum 1000 times faster than the existing techniques. We also performed imaging studies of single cTnT molecules and their motion in serum in real time and demonstrated the capability of this technique to measure ultra-low, clinically significant cTnT concentrations of 1 pg/mL.
Session 4: Superresolution Microscopy/Nanoscopy I
23 January 2022 • 9:30 AM - 10:30 AM PST | Room 201 (Level 2 South)
Session Chair: Mike Heilemann, Goethe-Univ. Frankfurt am Main (Germany)
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Author(s): Peter W. Tinning, Mark Donnachie, Jay L. Christopher, Deepak Uttamchandani, Ralf Bauer, Univ. of Strathclyde (United Kingdom)
23 January 2022 • 9:10 AM - 9:30 AM PST | Room 201 (Level 2 South)
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A novel SIM implementation using 2 micro-electro-mechanical system (MEMS) micromirrors is presented, allowing flexible control of the excitation illumination with reduced spatial footprint and cost-efficient integration. The 2D SIM approach utilizes two three-axis micromirrors with static positioning and piston control, which enable precise angular, radial and phase positioning of two interference beams in the back-aperture of a microscope objective. Isotropic 2D resolution enhancement can be achieved while simultaneously allowing spatial frequency control of the interference pattern to circumvent the missing cone problem in static 2D SIM.
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Author(s): Fei-Hung Chu, Gennady A. Smolyakov, Kevin J. Malloy, Alexander A. Ukhanov, Actoprobe LLC (United States)
23 January 2022 • 9:30 AM - 9:50 AM PST | Room 201 (Level 2 South)
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We have developed a novel class of AFM probes, Ultrafast Pulsed AFM Active Optical Probe (UFP AAOP), based on a mode-locked quantum-dot laser monolithically integrated into a cantilevered probe fabricated from GaAs, with a nano sized opening at the tip apex as the output aperture. The UFP AAOP is expected to provide pulses as short as ~ 0.1 - 5 ps and spatial optical resolution of ~ 50 - 200 nm. These unique optical probes will perform the functions of conventional AFM probes and provide both space- and time-resolved information about specimen’s chemical properties at the nanoscale simultaneously.
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Author(s): Carolin Grandy, Fabian Port, Univ. Ulm (Germany); Jonas Pfeil, Ulm University (Germany); Kay-Eberhard Gottschalk, Univ. Ulm (Germany)
23 January 2022 • 10:10 AM - 10:30 AM PST | Room 201 (Level 2 South)
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Cells adapt their actin cytoskeletons architecture to structural cues of the environment in all three dimensions. Nevertheless, how manipulating cell shape influences the actin cytoskeletons z-dimension is unstudied, but crucial for an understanding of the mutual influence of cell shape, cell tension and actin architecture. To study the effect of shape on the z-dimension of the actin cytoskeleton we combine metal-induced energy transfer as a super-resolution technique with micropatterning. This allows us not only to precisely manipulate the shape of the cell but also to regulate forces by changing the shape while studying specific actin structures with super-resolution.
Break
Coffee Break 10:30 AM - 11:00 AM
Session 5: Multimodal and Correlative Technologies
23 January 2022 • 11:00 AM - 12:10 PM PST | Room 201 (Level 2 South)
Session Chair: Linnea Olofsson, PicoQuant Photonics North America, Inc. (United States)
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Author(s): Peter Dahlberg, Stanford Univ. (United States)
23 January 2022 • 11:00 AM - 11:30 AM PST | Room 201 (Level 2 South)
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Identifying biomolecules of interest in cryogenic electron tomography (CET) reconstructions is made challenging by the lack of non-perturbative and specific labeling methods compatible with CET. Combining fluorescence and CET is a promising approach to overcome this labeling limitation. However, diffraction-limited fluorescence data has insufficient resolution to provide clear labeling in the crowded cellular environment. Super-resolution fluorescence techniques achieve resolution on the tens of nanometers scale, making them compatible with the length scales of interest in labeling biomolecules in CET. I will discuss the development of a cryogenic single-molecule based super-resolution imaging approach that achieves an average localization precision of less than ten nanometers and is compatible with the latest CET methods.
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Author(s): Fabian Port, Carolin Grandy, Jonas Pfeil, Kay-Eberhard Gottschalk, Univ. Ulm (Germany)
23 January 2022 • 11:30 AM - 11:50 AM PST | Room 201 (Level 2 South)
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Focal adhesions function as anchoring points to the extracellular matrix, yet also enable cells to sense and exert forces on their environment. Despite the important role of the focal adhesion complex in cellular adhesion, its structure and mechanoresponse remain difficult to resolve. Knowing the exact position of the proteins in the focal adhesion complex under strain is necessary to understand their working principle. We couple AFM techniques with Metal Induced Energy Transfer (MIET) to resolve positions at the nanoscale level. Here we show an initial analysis of the interplay between focal adhesion associated actin and force.
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Author(s): Kay Eberhard Gottschalk, Fabian Port, Carolin Grandy, Univ. Ulm (Germany); Jonas Pfeil, Ulm University (Germany)
23 January 2022 • 11:50 AM - 12:10 PM PST | Room 201 (Level 2 South)
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Cells adapt their actin cytoskeletons architecture to structural and mechanical cues of the environment. Focal adhesions function as anchoring points of actin to the extracellular matrix. How manipulating cell shape or exerting force on individual focal adhesions influences the actin cytoskeletons z dimension is unstudied. Metal induced energy transfer (MIET) is ideally suited to resolve actin structures in nm resolution in the z dimension . Therefore, we combined MIET with both micropatterning and high-resolution AFM measurements to study the influence of shape or force on actin structure. We show that these combinations reveals how focal adhesions adapt to external stimuli.
Break
Lunch Break 12:10 PM - 1:45 PM
Session 6: Superresolution Microscopy/Nanoscopy II
23 January 2022 • 1:45 PM - 3:15 PM PST | Room 201 (Level 2 South)
Session Chair: Ingo Gregor, Georg-August-Univ. Göttingen (Germany)
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Author(s): Mike Heilemann, Goethe-Univ. Frankfurt am Main (Germany)
23 January 2022 • 1:45 PM - 2:15 PM PST | Room 201 (Level 2 South)
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We present fluorophore labels that transiently and repetitively bind to their targets as probes for various types of super-resolution fluorescence microscopy. Such labels show a weak (~10 µM – 100 nM) affinity to a target and are kept in an imaging buffer that constitutes a reservoir with a high concentration of intact probes, enabling repetitive binding to the same target (we refer to these labels as “exchangeable labels”). This dynamic labeling approach minimizes photobleaching and yields a constant fluorescence signal over time, which has been beneficially exploited in SMLM [1-4], STED [5, 6], and super-resolution optical fluctuation imaging (SOFI) [7]. Multi-color, 3D, and live cell imaging, as well as imaging of large fields of view, is facilitated [4]. We further present the implementation of neural networks for multi-emitter localization to achieve multi-color SMLM with short acquisition times of one minute [8].
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Author(s): Jinhan Ren, Kyu Young Han, CREOL, The College of Optics and Photonics, Univ. of Central Florida (United States)
23 January 2022 • 2:15 PM - 2:35 PM PST | Room 201 (Level 2 South)
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Author(s): Abderrahim Boualam, Christopher J. Rowlands, Imperial College London (United Kingdom)
23 January 2022 • 2:35 PM - 2:55 PM PST | Room 201 (Level 2 South)
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Structured Illumination Microscopy (SIM) is a widely used super-resolution microscopy method, capable of imaging at twice the diffraction limit of conventional widefield microscopes. We developed a new method to assess in silico the spatio-temporal resolution limits of SIM, and demonstrated that its capacity to reconstruct super-resolved information is substantially worse than the time required to acquire a full stack of raw frames. We also applied our method to gauge the efficacy of a reconstruction method termed “rolling SIM” which claimed to improve the temporal resolution of SIM, and we showed that this is not the case.
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Author(s): Vahid Ebrahimi, Kyu Young Han, Univ. of Central Florida (United States); Jiah Kim, Univ. of Illinois (United States)
23 January 2022 • 2:55 PM - 3:15 PM PST | Room 201 (Level 2 South)
Session 7: Young Investigator Award Presentation
23 January 2022 • 3:15 PM - 3:35 PM PST | Room 201 (Level 2 South)
Young Investigator Award Presented by Ingo Gregor
Session PSun: Posters-Sunday
23 January 2022 • 5:30 PM - 7:00 PM PST | Moscone West, Lobby (Level 3)
Conference attendees are invited to attend the Sunday BiOS poster session. Come view the posters, enjoy light refreshments, ask questions, and network with colleagues in your field.

Poster Setup: Sunday 12:00 PM – 5:00 PM
View poster presentation guidelines and set-up instructions at:
https://spie.org/PW/Poster-Guidelines
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Author(s): Jongwu Kim, Dug Young Kim, Yonsei Univ. (Korea, Republic of)
23 January 2022 • 5:30 PM - 7:00 PM PST | Moscone West, Lobby (Level 3)
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In this presentation, we will introduce a modified blaze condition of a digital micromirror device for structured illumination microscopy. This condition aims to remove special mask at the Fourier plane, which blocks unwanted orders to make two beams or three beams interference. Proposed alignment is intrinsically free from zeroth order of diffraction light. So we can generate structured light by uploading proper patterns onto DMD with an iris diaphragm instead of special mask. Adjusting the size of the iris diaphragm, structured light with various periods can be illuminated to the sample. The basic concept, proper pattern making method and experiment results with a modified blaze condition will be presented.
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Author(s): Arpan Dey, Sudipta Maiti, Tata Institute of Fundamental Research (India)
23 January 2022 • 5:30 PM - 7:00 PM PST | Moscone West, Lobby (Level 3)
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The “which oligomer” question in the field of amyloid-linked diseases asks which oligomer, of many types coexisting in equilibrium, is toxic. Here we describe a group of new single molecule photo-bleaching techniques which can provide an oligomer-stoichiometry-specific understanding of their properties. One measures the affinity of individual types of oligomers to plasma-membrane mimicking lipid bilayers. Another [termed “Q-SLIP” for Quencher induced Step Length Increase in Photobleaching) measures solvent exposure of individual oligomers. Using Q-SLIP, we observe that amylin oligomers with specific stoichiometries have remarkably different properties compared to the other stoichiometries.
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Author(s): Dongeun Kim, Wonsang Hwang, Yonsei Univ. (Korea, Republic of); Sucbei Moon, Kookmin Univ. (Korea, Republic of); Dug Young Kim, Yonsei Univ. (Korea, Republic of)
23 January 2022 • 5:30 PM - 7:00 PM PST | Moscone West, Lobby (Level 3)
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Time-correlated single photon counting (TCSPC) is a standard measurement method for time-resolved spectroscopy, but suffers with pile-up effect in photon acquisition. To avoiding the pile-up effect, conventional TCSPC is usually operated at low count rate (<10%). Various methods have been suggested to overcome this limit. Digital TCSPC is recently suggested technique, which implements core electronics of TCSPC as digital process. Based on pulse discrimination, digital TCSPC can distinguish single-photon and multi-photon arrival events, therefore can achieve pile-up-free counts of photon. The maximum count rate of digital TCSPC depends on efficiency of the pulse discrimination. In this study, we verified gain variance affects the efficiency of the pulse discrimination, and how the count rate can be achieved in digital TCSPC.
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Author(s): Kristian Lauritsen, Dietmar Klemme, Guillaume Delpont, Evangelos Sisamakis, Marcelle König, Uwe Ortmann, Felix Koberling, Rainer Erdmann, PicoQuant GmbH (Germany)
23 January 2022 • 5:30 PM - 7:00 PM PST | Moscone West, Lobby (Level 3)
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Pulsed Interleaved Excitation (PIE) is a well-known technique for synchronizing and/or delaying pulsed lasers to alternately excite donor- and acceptor-labeled species within single-molecule FRET (smFRET) studies. Laser pulses are separated on a nanosecond (ns) time scale to allow for the simultaneous recording of the temporal behavior of the two species that are excited by the corresponding laser. This allows to differentiate between a FRET pair - even with very low energy transfer efficiency - from a molecule with either absent or non-fluorescing acceptor. Alternating-Laser EXcitation (ALEX) is another approach for smFRET studies which is based on interleaved continous wave (cw) laser excitation on the microsecond (μs) time scale. Here, we present the combination of both techniques (dubbed PIE-ALEX) in order to optimize excitation patterns within smFRET studies. A new generation of picosecond (ps) pulsed and fast cw-switched laser heads enables a simultaneous PIE and ALEX excitation scheme. MHz repetition rates are used for pulsed excitation for the donor, whereas fast-switched cw excitation in the range of 100 ns is used for the excitation of the acceptor species.
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Author(s): Ingo Gregor, Niels Rademacher, Georg-August-Univ. Göttingen (Germany); Max Tillmann, Matthias Patting, PicoQuant GmbH (Germany); Jörg Enderlein, Georg-August-Univ. Göttingen (Germany); Felix Koberling, PicoQuant GmbH (Germany)
23 January 2022 • 5:30 PM - 7:00 PM PST | Moscone West, Lobby (Level 3)
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Simultaneous investigations of the spatial arrangement and relative composition of proteins are of crucial importance in live-cell imaging. It was recently shown that Image Scanning Microscopy (ISM) can double the confocal resolution. Additionally, fluorescence lifetime based imaging (FLIM) opens an additional dimension in multispecies identification and separation. We combine these two techniques using an array of single-photon detectors together with a multichannel TCSPC and time tagging system for time-resolved single photon detection via tens of channels. First results illustrate the spatial resolution improvement and the selection of suitable fluorescent dyes to allow for lifetime based multiplexing.
Conference Chair
Georg-August-Univ. Göttingen (Germany)
Conference Chair
PicoQuant GmbH (Germany)
Conference Chair
PicoQuant GmbH Berlin (Germany)
Program Committee
The Univ. of Southern California (United States)
Program Committee
Friedrich-Schiller-Univ. Jena (Germany)
Program Committee
Univ. of Oxford (United Kingdom), Friedrich-Schiller Univ. Jena (Germany)
Program Committee
Georg-August-Univ. Göttingen (Germany)
Program Committee
Imperial College London (United Kingdom)
Program Committee
Ewa M. Goldys
The Univ. of New South Wales (Australia)
Program Committee
Univ. of North Texas Health Science Ctr. at Fort Worth (United States), Texas Christian Univ. at Fort Worth (United States)
Program Committee
Goethe-Univ. Frankfurt am Main (Germany)
Program Committee
KU Leuven (Belgium)
Program Committee
Zhen-Li Huang
Huazhong Univ. of Science and Technology (China)
Program Committee
Univ. Bielefeld (Germany)
Program Committee
Univ. of California, Los Angeles (United States)
Program Committee
Institute of Chemistry (China)