Research.jefferson.edu
MitoCare Service Centers: (1) Confocal, Multiphoton and Super-resolution Imaging, (2) Electron Microscopy & Tomography, (3) Seahorse XF24
(1) Confocal, Multiphoton and Super-resolution Imaging
Director – Gyorgy Hajnoczky, M.D. Ph.D. & David Weaver
Contact – David.Weaver@jefferson.edu
215-503-0169
To schedule this service: iLabs
|Mission, Goals, Capabilities |The confocal, multiphoton and super-resolution imaging facility will provide |
| |services particularly focused on high resolution fluorescence visualization. The |
| |available confocal/multiphoton system provides for a range of experiments from |
| |sub-cellular localization to mapping of large tissue sections. The |
| |super-resolution capability utilizes single molecule localization by stochastic |
| |imaging (StORM/PALM) to circumvent the diffraction limit of optical microscopy and|
| |provide resolution at < 50 nm. The facility will provide access to these |
| |instruments for researchers, training on their use and consultation on |
| |experimental design and analysis. |
|Major Equipment |Zeiss LSM780 NLO confocal/multiphoton microscope: 405, 457, 488, 514, 561, 594 and|
| |633 nm laser lines for single photon excitation. Chameleon Vision II system |
| |(Coherent) for MP illumination. The confocal scanner is coupled to a Zeiss |
| |inverted microscope (Axio Observer). Three internal detectors including Meta |
| |detector for spectral imaging. Two external, non-descanned detectors for |
| |multi-photon imaging. Transmitted light detector for DIC/phase contrast. 10x, 20x,|
| |25x, 40x, 63x and 100x objectives. Targeted illumination capability for |
| |photo-bleaching/activation; motorized focus for z-stacks; encoded, motorized |
| |stage; temperature and humidity controlled incubator with CO2 regulation. Zen |
| |acquisition and analysis software. |
| |Bruker Vutara 352 super-resolution microscope: StORM/PALM type super-resolution |
| |with 488, 561, 639 and 750nm lasers for excitation and 405nm for photoswitching. |
| |Detection via Quad-Field projection onto a sCMOS camera (Hamamatsu Orca-Flash4.0 |
| |V2) allows simultaneous imaging of two colors in two focal planes or up to four |
| |colors in a single-plane. Includes a motorized TIRF module and |
| |temperature-controlled incubation for live cell imaging. Includes software for |
| |analysis of single-molecule localization and quantitative analysis of |
| |super-resolution experiments and/or single particle tracking. |
|Services: |High resolution, 3D imaging of live and fixed specimens |
| |Tile-scan images for large areas |
| |Imaging in multi-well formats |
| |Photo-activation, bleaching or other optogenetic techniques |
| |Tissue imaging with multi-photon excitation |
| |Sub-diffraction-limit molecular localization microscopy. |
| |Training on microscope use and data analysis |
| |Consultation on experiment design. |
(2) Electron Microscopy & Tomography
Director: Gyorgy Csordas, MD. - 215-955-4556
Contact: TBN Tech
|gyorgy.csordas@jefferson.edu |
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|To schedule this service: iLabs |
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|Mission, Goals, Capabilities |The Research Electron Microscopy Laboratory in the Department of Pathology, |
| |Anatomy and Cell Biology aims to provide researchers with ultrastructure in |
| |specimen prepared from tissue, cellular or subcellular preparations. We are also |
| |open (on case by case base) to collaborative consultations to assist users in TEM|
| |related experimental design and analysis. Current capabilities allow 2D |
| |transmission electron microscopy. Tilt-angle tomography of ~150 nm thick sections|
| |of resin-embedded samples (or negative-stained particles) will be available* for |
| |limited collaborations. |
|Major Equipment |The core instrument is an FEI Tecnai 12 120 keV digital TEM, equipped with an AMT|
| |XR111 CCD camera and upgraded with a tomography package that uses the FEI Xplore|
| |3D Tomography software. Addition of a high field of view tomography holder and |
| |Xplore3D retrofit package allows acquisition of electron tomograms at high tilt |
| |angles. The sample preparation laboratory has two Leica UCT ultramicrotomes. |
|Services: |Services available include: |
| |I. (20% technician/operator effort, ~1 day scope time/week). Providing training |
| |and supervision for user to obtain images with the scope. Providing |
| |instructions/demo for routine and non-routine fixation and embedding. Compact |
| |scoping assignments. |
| |II. (40% technician/operator effort ~2 days scope time/week) in addition to I, |
| |extended limited scoping assignments, electron tomography (limited |
| |collaborations). Resin embedding. Ultramicrotomy and (post)staining of thin and |
| |thick sections. |
*: Tilt-angle tomography requires a currently missing part, the tomography sample holder, for which a capital budget request is submitted.
(3) Seahorse XF24
Erin Seifert, PhD
Erin.Seifert@jefferson.edu
To schedule this service: iLabs
|Mission, Goals, Capabilities |The Seahorse XF24 instrument is a sensitive, high-throughput (24-well) |
| |easy-to-use technology to measure changes in O2 tension and proton concentration |
| |in the medium, as a function of time (~10 second resolution). The typical |
| |application is to interrogate mitochondrial bioenergetics in adherent |
| |preparations (intact or permeabilized adherent cells, or isolated mitochondria |
| |electrostatically adhered to plastic). The technology is endorsed by the |
| |mitochondrial biology community, and has been used in 100s of publications in |
| |multiple areas of biology). |
| |The service center will 1) provide access to the Seahorse XF24, 2) provide basic |
| |training, and 3) ensure the functionality of the XF24. |
| |The XF24 is located within the MitoCare Center, JAH 528, at the “Seifert lab” |
| |space. |
|Major Equipment |Seahorse XF24 Instrument; PC with acquisition software and Excel for initial |
| |analysis of data; 37ºC, ambient CO2 incubator. |
|Services |To provide access to the Seahorse instrument, as well as to provide 1) training |
| |for 1st time users in instrument use and navigating the data output file, and |
| |refresher training if needed for returning users, 2) basic information about cell|
| |seeding density, concentrations of uncoupler and substrate conditions, 3) access |
| |to an incubator for overnight hydration of the cartridge, 4) access to bench |
| |space for plate preparation, 5) access to Excel for initial data analysis, 6) |
| |oversight of instrument scheduling, 7) leadership to ensure the functionality of |
| |the instrument, by regular inspection of instrument function, by arranging the |
| |purchase of the service contract, scheduling annual on-sight preventative |
| |maintenance and any additional maintenance. |
| |Not provided: consumables (e.g. custom culture plates, cartridges, assay media, |
| |permeabilizing reagents, cartridge hydration solution), pipettors, assistance |
| |with experimental design and data interpretation, anything related to and |
| |including an actual seahorse. |
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