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Cell & Molecular Imaging Shared Resource

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Presentation on theme: "Cell & Molecular Imaging Shared Resource"— Presentation transcript:

1 Cell & Molecular Imaging Shared Resource
SHARED RESOURCES Cell & Molecular Imaging Cell & Molecular Imaging Shared Resource SCTR Scientific Retreat on Aging Related Research MUSC, October 20, 2017 Monika Gooz, M.D., Ph.D. Manager, Cell & Molecular Imaging Core

2 CMI Overview Cell & Molecular Imaging: Overview
SHARED RESOURCES Cell & Molecular Imaging: Overview CMI Overview John J. Lemasters, MD, PhD, Director Anna-Liisa Nieminen, PhD, Co-Director Monika Beck Gooz, MD, PhD, Facility Manager Li Li, Ph.D, Imaging Specialist

3 CMI Overview Cell & Molecular Imaging: Overview
SHARED RESOURCES Cell & Molecular Imaging: Overview CMI Overview Specific objectives of the HCC CMI Shared Resource are to: 1) Give investigators access to state-of-the-art microscopic techniques; 2) Conduct training in confocal and multiphoton microscopy (independent usage mostly; assisted imaging available) 3) Provide consultation and assistance concerning: experimental design, sample preparation, probe selection data analysis for confocal and multiphoton microscopy applications, assistance in preparation of high quality images for publications and grant applications; 4) Establish state-of-the-art applications in the CMI Shared Resource that address the emerging needs of the members of HCC and MUSC; 5) Organize microscopy workshops and seminars to educate MUSC researchers about microscopy techniques for their research.

4 CMI Overview Cell & Molecular Imaging: Equipment
SHARED RESOURCES Cell & Molecular Imaging: Equipment CMI Overview The CMI Shared Resource houses the following equipment: Zeiss LSM 880 Quasar NLO multiphoton/confocal system with Airyscan super-resolution detector (DD521) Olympus Fluoview FV1200 silicon oil-immersion intravital multiphoton microscope (DD521) Olympus Fluoview FV 10i LIV water-immersion confocal microscope (DD521) BD BioSciences CARV II disk-scanning confocal system (DD526- dark room) Zeiss LSM 510 META confocal microscope (HCC355) Imaging workstations running Metamorph, ImageJ FIJI, Photoshop, Duolink ImageTool and other software for offline image processing and analysis (DD521)

5 CMI Overview Cell & Molecular Imaging: Equipment
SHARED RESOURCES Cell & Molecular Imaging: Equipment CMI Overview Zeiss LSM 880 Quasar NLO multiphoton/confocal system (2015, SIG) with Airyscan super-res detector (2016, JJL grant funds) Airyscan Super-Resolution: 140 nm (xy) Lasers: 405, 488/514, 561 nm and 633 nm and a tunable Ti-Sapphire multiphoton laser Environmental control for live cell imaging applications (receptor internalization, mitochondrial metabolism, cell death, ROS production in real-time) Zen software for image acquisition and analysis (co-localization, FRET, FRAP) 34-Channel Quasar spectral detection Carl Zeiss Microscopy

6 Cell & Molecular Imaging: Equipment
SHARED RESOURCES Cell & Molecular Imaging: Equipment Airyscan image examples Figure 1: Airyscan of mitochondria of a HepG2 hepatoma cell labeled with Mito-SOX Red and Mitoview 633 (pseudocolored green). Note that spots of red/yellow MitoSOX Red reside within the green-labeled mitochondrial matrix. Bar is 2 µm. Figure 2: Airyscan of HepG2 cell labeled with PicoGreen (DNA) and TMRM (ΔΨ). Dotted line shows perinuclear area with low ΔΨ mitochondria. Figure 3: VDAC1/2 knockdown inhibits mitochondrial fission. HepG2 cells were loaded with TMRM. Images of Drs. John J. Lemasters and Eduardo N. Maldonado

7 CMI Overview Cell & Molecular Imaging: Equipment
SHARED RESOURCES Cell & Molecular Imaging: Equipment CMI Overview Olympus Fluoview FV1200 intravital multiphoton microscope Ti-Sapphire multiphoton tunable laser Thick section and deep tissue imaging (brain, liver, kidney) Second harmonic imaging (collagen, muscle fiber) Intravital ROS detection in mouse liver (Jiangting Hu, Lemasters lab)

8 CMI Overview Cell & Molecular Imaging: Equipment
SHARED RESOURCES Cell & Molecular Imaging: Equipment CMI Overview Olympus Fluoview FV 10i LIV confocal microscope Easy to use for first time users Automated imaging Environmental control, CO2/hypoxia for live cell imaging application Fluoview software for imaging and image analysis

9 CMI Overview Cell & Molecular Imaging: Equipment
SHARED RESOURCES Cell & Molecular Imaging: Equipment CMI Overview BD BioSciences CARV II disk-scanning confocal system Spinning disk fluorescence confocal imaging Zeiss Axiovert 200 M for brightfield and widefield fluorescence microscopy H&E, Masson staining, BrdU incorporation Polarization microscopy (collagen)

10 C&MI: 2017 SIG – resubmission
SHARED RESOURCES C&MI: 2017 SIG – resubmission Quantitative single-molecule localization (dSTORM and PALM) 3D super-resolution imaging to depths into tissue of up to 20 m using biplane technology Resolution: 20 nm XY, 40 nm Z (up to 10-times greater than our existing microscopes) 2-Color simultaneous imaging Lab of Hainan Lang, unpublished

11 C&MI: 2017 SIG – resubmission
SHARED RESOURCES C&MI: 2017 SIG – resubmission Applications Protein localization Co-localization studies (proximity) Membrane structures Exosomes Cytoskeleton Infectious disease - bacteria Zebrafish embryo (development) Receptor trafficking Robert Gemmill, Ph.D. (MUSC)

12 C&MI: Need assessment – Vectra Polaris
SHARED RESOURCES C&MI: Need assessment – Vectra Polaris FFPE and tissue microarray Multispectral imaging: up to 7 “colors” Can image, analyze, and quantify fluorescence and brightfield (H&E and Trichrome) staining High speed digital whole slide scanning inForm software analyzes cell types (phenotyping) and tissue types Application: Tumor and tumor microenvironment Cancer Immunology Polaris, Perkin Elmer

13 CMI Overview Cell & Molecular Imaging: Education
SHARED RESOURCES Cell & Molecular Imaging: Education CMI Overview Semi-annual Charleston Workshop on Light Microscopy for the Biosciences June 4-9, 2017 Maximum 24 participants (technical personnel, graduate students, post-docs, PIs) One graduate credit for participation is also available to students at MUSC and the College of Charleston (Pharmacy 722) Lecture topics cover basic principles of microscopy and advance to contemporary high-end confocal/multiphoton imaging Keynote Speaker: Joerg Bewersdorf (Yale University) Topic: Super-resolution microscopy Website is currently in development to advertise the course better in the future Fee is $650 Course is supported by HCC with 5 full scholarships Supported by vendors: Carl Zeiss Microscopy LLC, Leica, Olympus, Nikon, ThermoFisher, ibidi Gooz Shared Resource Report May 5, 2017

14 CMI Overview Cell & Molecular Imaging: Education CMI Seminar series:
SHARED RESOURCES Cell & Molecular Imaging: Education CMI Overview CMI Seminar series: Spring 2016 Super-resolution microscopy “TIRF technology” (John Marchlenski, Charles Reichley, Matt Maudsley Olympus) “Quantitative Super-Resolution Applications in Life Science Using Single Molecule Localization Microscopy” (Manasa Gudheti, Vutara) “Airyscan super-resolution microscopy” (Rob Mihalik, Zeiss) “Super-resolution imaging beyond the diffraction limit” (Nick Villani, Nikon) January 19, 2017 “Navigating the immunological microenvironment in situ by automated multiplexed immunohistochemical imaging” presented by Perkin Elmer’s application scientist Edward Stack, Ph.D. April 26, 2017 “Advances in 3D single molecule super-resolution imaging using the Bruker Vutara 352 Microscope" by Manasa Gudheti, Ph.D. Websites academicdepartments.musc.edu/ccdir

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