High Spatial and Temporal Resolution Imaging Core Services and Equipment

HSTRI Core Services and Equipment

The High Spatial and Temporal Resolution Imaging (HSTRI) Core at the University of Nevada, Reno offers a comprehensive range of cutting-edge microscopy systems and imaging analysis tools to support a wide variety of biomedical research needs.

Services

Below are the hourly rates for the HSTRI microscope systems:

*Off Peak Hourly Rate: This discounted rate will apply outside of the peak Monday through Friday, 8 a.m. to 5 p.m. business hours. The discount results from having no core staff available.

Leica Stellaris 8 STED Super-Resolution Microscope

• Leica DMi8 inverted fluorescence microscope equipped with Adaptive Focus Control and Closed Loop Focus (20 nm re-positioning accuracy)
• LED light engine for eyepiece visualization with filter cubes (DAPI, CFP, GFP, Rhod LP) and a scanning XY Stage.
• Assay Editor for multiwell plate screening
• Super Z high-speed / high precision Z galvo stage for XYZ, XYZT, XZY, and XZYT high-speed imaging.
• 4 objectives
  • 40x, NA 1.30, Plan APO (Oil) – 0.17mm FWD
  • 86x, NA 1.20, Plan APO motorized correction collar STED (Water) – 0.3mm FWD
  • 93x, NA 1.30, Plan APO STED motorized correction collar (Glycerol) – 0.3mm FWD
  • 100x, NA 1.40 Plan APO STED (Oil) - 0.13mm FWD
• Lasers
  • White Light Supercontinuum Laser Extreme (440 nm-790 nm)
  • 405 nm Diode - 50 mW CW
• High performance 8 channel crystal Based Acousto optical beam splitter with 10 µs switch time. · A Tandem Scanner FOV + 8K Resonant Point Scanner with prism-based spectral imaging (extended red range 410 nm - 850 nm).
• Light detection is accomplished via Power HyD Detectors:
  • 2 Spectral HyD S (high QE/PDU, highest dynamic range, high light level OK, photon counting, spectral, FLIM)
  • 2 Spectral HyD X (high QE/PDU, high dynamic range, photon counting, spectral, ideal for FLIM)
  • 1 Spectral HyD R (red optimized high QE/PDE 720 nm - 850 nm, high dynamic range, photon counting, spectral, ideal for FLIM)
  • PMT based brightfield detector.
• Lightning Mode (spectral, multi-channel super-resolution imaging: 120 nm XY & ~200 nm Z resolution at 1.4 NA; works with all detectors and all objectives)
• 3D Tau STED nanoscopy with 592 nm / 660 nm / 775 nm depletion lasers (86x, 93x & 100x STED WHITE objectives (sub 50 nm XY resolution; sub 130 nm Z resolution; multi-channel, multi- color standard, gated or Tau STED)
• Spectral Fluorescence Lifetime Imaging (FLIM; FALCON Fast FLIM, FLIM FRET, Phasor Analysis; Tau sense average photon arrival time with gating, contrast, and separation).
• The system is controlled by LAS X Software running on a high-performance HP Z6G4 computer workstation comprising 2 HD monitors, 192 GB RAM / 6 TB HDD storage, Nvidia Quadro RTX 6000 graphics with 24 GB memory, 4608 CUDA cores, and 2 Thunderbolt ports (TB3).
• OKO Labs opaque microscope enclosure environmental control with active mixing CO2, Humidity, and Heat.

Leica Stellaris 5 Confocal Microscope

• Leica DMi8 inverted automated fluorescence microscope.
• 5 objectives
  • 5x, NA 0.15, HC PL FLUOTAR (Air)
  • 10x, NA 0.40, HC PL APO CS2 (Air)
  • 20x, NA 0.75, HC PL APO CS2 (Air)
  • 40x, NA 1.30, HC PL APO CS2 (Oil)
  • 63x, NA 1.40, HC PL APO CS2 (Oil)
• 6 solid-state diode lasers for multicolor imaging
  • 405 nm
  • 448 nm
  • 488 nm
  • 514 nm
  • 561 nm
  • 638 nm
• Fast laser scanning head (tunable speed large FOV 22 mm linear, cooled point scanner)
• Motorized DIC and phase contrast
• 4 Peltier-cooled SuperResolution HyD S High PDE SiPM Detectors allowing for true photon counting, power counting & analog modes, and BF imaging.
• Adaptive Focus Control with 20 nm repositioning accuracy
• High temporal (131 FPS, 512 x 16 x 4 Spectral + 1 BF for 655 FPS total) and spatial (8K x 8K) resolution
• LAS X Software controls this instrument and is capable of 3D GPU-based visualization, online linear unmixing, live data mode, FRET, FRAP, and FLIP imaging modes.
• High-end Windows 10-based computer (HP CUDA Workstation) and a 38" high-resolution curved monitor

Leica Stellaris DIVE Multiphoton Microscope

• The Stellaris DIVE Multiphoton facilitates multicolor, deep tissue imaging of 1mm or beyond.
• Upright Dm6 microscope
• 8K tandem scanner
• 3 hybrid detectors
• 4 standard laser lines (405, 488, 561, 638nm)
• Long working distance objectives
  • 10x, NA 0.4, HC PL APO CS2 (Air)
  • 20x, NA 0.75, HC PL APO CS2 (Air)
  • 10x, NA 0.3, HC APO W U-V-I (Water)
  • 25x, NA 0.95, HC FLUOTAR W VISIR (Water)
• Coherent Chameleon Vision ll multiphoton laser (600-1080nm).
• 4Tune non-descanned detector system, equipped with 2 detectors.
• 4 channels can be acquired simultaneously, or an unlimited number when imaging sequentially.
• VarioBeam Expander (VBE) - combines tunable beam diameter and tunable divergence. This offers maximum depth, best resolution, and full-color correction.
• Multiphoton imaging and lifetime information can be combined using FLIM.

Olympus-Based Spinning Disk Confocal and TIRF Microscope

• Olympus IX83 microscope
• 4 objectives
  • 20x, NA 0.45, LUC PlanFl N (Air)
  • 40x, NA 0.55, SLCPlanFl (Air)
  • 60x, NA 1.49, Apo N (Oil)
  • 100x, NA 1.50, UPlanApo (Oil)
• Automated XY stage for multi-position acquisition
• TrueFocus Zero drift automatic focus
• Andor Integrated Laser Engine (ILE) and Borealis with 405-, 488-, 538-, and 640-nm solid-state lasers and integrated acousto-optic tunable filter (AOTF)
• The spinning disc confocal side consists of a Yokogawa CSU-21 Nipkow spinning-disk confocal scanner unit and an iXon X3 cooled EMCCD camera.
• The TIRF side contains a 2-line motorized Olympus IX3 CellTIRF Illuminator and an iXon Ultra cooled EMCCD camera.
• Seamless switching between confocal and TIRF imaging modalities
• Oko-Touch stage-top incubator provides temperature and CO2 regulation for long-term imaging of live cells
• CellSens Dimension software running on a Windows 10-based PC

Olympus TIRF system with FRET and patch clamp

• Olympus IX83 epifluorescence microscope
• TrueFocus Zero drift automatic focus
• Olympus CellTIRF module for TIRF microscopy
• 4 objectives
  • 20x, NA 0.45, LUC PlanFl N (Air)
  • 40x, NA 0.55, SLCPlanFl (Oil)
  • 60x, NA 1.45, Apo N (Oil)
  • 100x, NA 1.49, UPlanApo (Oil)
• Hamamatsu ImagEM EMCCD digital video camera (C9100-13): refresh rate ≈ 32 Hz at full 512 x 512 pixels without binning.
• Vortran STRADUS® VERSALASE laser launch housing three TTL-shutter-controlled diode lasers (50 mW 488 nm, 50 mW 561 nm and 140 mV 637 nm).
• TTL-shutter-controlled Coherent 40 mV 445 nm laser.
• Photometrics DV2 Dual View beam splitter allows CFP/YFP FRET imaging in either standard epifluorescence or TIRF imaging modes.
• HP SB EliteDesk 800 computer running Windows 10.
• Visiview software (Visitron Systems) for imaging and microscope control, and Olympus CellTIRF (v. 1.4) for control of the TIRF module.
• The microscope is also coupled to a patch clamp workstation.
  • Axopatch 200A patch clamp amplifier (Axon Instruments, Inc.)
  • MHW-3 Narishige micromanipulator
  • TMC (series 63-543) vibration-free isolation table
  • TMC Faraday cage o GWINSTEK digital storage oscilloscope (model GDS-2102)
  • Digidata 1322A acquisition system
  • PClamp software (v. 9; Axon Instruments, Inc.) running on a separate Windows 7-based desktop computer
  • This configuration allows for simultaneous recording of membrane currents or voltage, and TIRFM/FRET imaging.
• State-of-the-art perfusion system to allow for rapid solution changes during physiological experiments

Leica GSD-3D Super-Resolution Microscope

• Inverted LeicaDMI6000B microscope
• Multiple imaging modes
  • GSD super-resolution
  • Fully automated TIRF (also available with GSD)
  • EPI fluorescence (also available with GSD)
  • Brightfield
  • DIC/PH
• Lateral resolution of 20 nm and axial resolution of 50 nm, providing advanced 3D localization of cellular structures
• Accurion Halcyonics i4 Vibration Isolation table to minimize drift
• Field of view
  • 18 x 18 μm (GSD high power mode)
  • 40 x 40 μm (GSD large field of view mode)
  • 50 x 50 μm (TIRF)
• 3 objectives
  • 10x, NA 0.30, HCX PL FLUOTAR (Air)
  • 100X, NA 1.47 (Oil)
  • 160X, NA 1.43, HC Pl Apo Corr GSD(Oil)
• 3 lasers (500mW) for multicolor imaging
  • 488 nm
  • 532 nm
  • 642 nm
• Andor iXon3 897 EMCCD camera
• Real-time image processing and display of results as they are acquired
• Image acquisition via LAS AF software running on an HP Z620 workstation

3i Lattice LightSheet Microscope

• The HSTRI Core manages a custom-built 3i Lattice LightSheet Microscope that is capable of 3D live cell imaging.
• The Lattice LightSheet is a system designed to push the spatial and temporal resolution limits in live cell imaging.
• Cylindrical lenses stretch and collimate the beam to form a sheet projected onto a spatial light modulator (SLM).
• The SLM generates an optical lattice of Bessel beams.
• An annular mask acts as a zero-order filter, removing artifacts and lengthening the sheet.
• Lightsheet thickness is 0.4 µm at 50 µm length.
• Galvos dither the sheet in X and sweep the sheet in Z.
• 25x/1.1NA 28x/0.71NA water immersion detection objectives
• Piezo x,y translation stage and piezo imaging objective control.
• Images are acquired using a high-speed high-resolution 2Kx2K Hamamatsu ORCA-Flash4.0 V3 sCMOS camera.
• Samples are excited with 405, 488, 560, and 642 nm lasers.
• Image acquisition and analysis are performed by 3i SlideBook software (v. 6) running under Windows 10 on a powerful desktop computer comprised of Dual 16-Core Xeon Gold 2.9GHz processors, 128GB RAM, 8GB NVIDIA Quadro RTX4000 workstation graphics card, 512GB OS SSD, 8TB Fast Acquisition Drive, and 20TB additional storage.
• The system is also capable of Structured Illumination Microscopy (SIM), which allows breaking of the diffraction limit for single molecule imaging and photomanipulation.

Olympus-based full-color imaging system

• Olympus IX 50 microscope
• Transmitted light for brightfield imaging
• Perfect for imaging of histology sections.
• EPI fluorescence
• Filter cubes for DAPI, TRITC and Texas Red.
• 6 objectives
  • 2x, NA 0.06, Plan N (Air)
  • 4x, NA 0.13, UPlanFl (Air)
  • 10x, NA 0.30, UPlanFl (Air)
  • 20x, NA 0.40, LCPlanFl Ph1 (Air)
  • 40x, NA 0.60, LCPlanFl Ph2 (Air)
  • 60x, NA 0.70, LCPlanFl (Air)
• Qimaging MicroPublisher 5.0 Digital CCD Color Camera
  • 5-million-pixel sensor; 2560 x 1920
  • 3.4μm x 3.4μm pixel size
  • Previewing and focusing in real time
  • Frame rate - 25fps (full field of view)

Imaris Software

• The HSTRI Core has one fixed and one floating license of the advanced imaging analysis software package, Imaris (v. 10.2; Oxford Instruments).
• Imaris is designed for motion detection (average and instantaneous speed, acceleration, track length, cell cycle duration), cell biology (cell volume, number of vesicles per cell, distance to membrane, vesicles per nucleus, distance to cell center, spot or particle detection (count, position (x, y, and z), distance between spots, distance to surface object, speed) and surface (area, volume, intensity, ellipticity, sphericity) analysis.
• AI machine learning is available for training the software to detect almost any object.
• The two licenses comprise several specialized Imaris packages designed for specific tasks: 1) Imaris XBT/Colocalization module, 2) Imaris ClearView Deconvolution module; and 3) Imaris Neuroscientist module.
• The fixed license is associated with a single high-end Windows 10-based computer workstation located in room 44 of the Savitt Medical Sciences building. PC specs include:
  • 16 x Total Xeon Scalable Processor Cores/2.1 GHz, 2 x Intel Xeon Scalable Processors, 32 Cores with hyperthreading enabled
  • 256 Gb high-performance 2933 MHz DDR4 ECC registered memory o 10.6+GBperProcessorCore
  • 2 x 480 GB solid-state SATA lll (SSD) Enterprise hard drive for operating system
  • 6 x 1 TB SATA lll 7200 RPM Enterprise hard drive for data storage
  • A GeForce RTX-2080Ti /11Gb/GDDR6 memory/graphics Card
  • 2 x 10/100/1000/10000 10 GigE SFP + network ports
  • The floating license can be accessed anywhere within the UNR Med by trained users, allowing them to analyze their images on their own computer.