Resources

Links

Microscope and workstation list for Porter Bioscience LMCF Detailed instrument descriptions Microscopy links & search engines Software download links Fluorescence & spectra viewers Help with sample preparation, image acquisition and analysis! Standard operating procedures Cell lines & fluorescent reporters
Microscope and workstation list for Porter Bioscience LMCF

Microscopes:

Nikon Ti2E AXR NSPARC laser scanning confocal.

Nikon TiE A1+/N-SIM laser scanning confocal with TIRF.  

Nikon TiE inverted spinning disk confocal.

Nikon TiU inverted widefield fluorescence and RGB.

Nikon SMZ745 zoom stereo.

Olympus FVMPE-RS twin laser 2-photon .

Olympus IX81 inverted widefield fluorescence.

Olympus MVX10 macrozoom fluorescence and RGB.

Yokogawa CV1000 inverted spinning disk confocal.

Zeiss Dicovery.V12 stereo fluorescence.

 

Workstations:

Hewlett Packard Z4 v7 with Nikon Elements AR image processing, measurement analysis engine, 3D analysis, spectral deconvolution, 2D/3D deconvolution, more.

Hewlett Packard Z8 G4 with Imaris 9.2.1 for Cell Biologists, Olympus cellSens with 3D deconvolution, FIJI, and Yokogawa CV1000 viewer.

Dell Precision 7910 with Nikon Elements AR with SIM reconstruction, MATLAB, Zeiss Zen Lite, Cell Profiler, FIJI, ICY, and Yokogawa CV1000 viewer.

Dell Precision T3500 with Yokogawa CV1000 viewer, 3i Slidebook, FIJI

 

https://www.colorado.edu/lmcf/choose-type-microscopy

2-photon microscopy:

‘GEMINI’ – Olympus FVMPE-RS twin laser 2-photon microscope.

This is a dedicated 2-photon microscope built on an Olympus BX63LF upright frame. The microscope is equipped with two separate, pre-chirped, tunable, pulsed IR lasers, a Spectra Physics Insight X3 (680-1300 nm) and Spectra Physics MaiTai eHP DeepSee (690-1040 nm). The microscope has four PMT detectors, two enhanced and two high-sensitivity GaAsP. All optics are designed for long wavelength transmittance and reflectance. The hybrid scanning system with galvanometer and resonance scanners is ideal for high-temporal resolution imaging. Available 2-photon optimized objectives include a 5x NA 0.10 air WD 20 mm, 10x NA 0.60 water immersion WD 3.3 mm, 25x water immersion NA 1.05 WD 2.0 mm and 60x water immersion NA 1.10 WD 1.5 mm. A 20x air, NA 0.45 WD 8.2 mm is available for GRIN lens work.

This system is controlled with advanced Olympus Fluoview acquisition software, a motorized nosepiece and X-Y Prior Z-Deck stage allowing for multipoint, tiling, Z-series time-lapse imaging, etc. Other features include auto-alignment of the lasers, beam- shaping for improved deep focus, and a motorized correction collar for the 25X objective to correct for refractive index over depth to provide maximum brightness, contrast and resolution through the entire imaged volume. Laser output power can also be software adjusted and tuned through the imaging depth.

A sample warming pad (Stoelting) and isoflurane anesthesia is available for imaging small animals. This microscope is housed in an independent, secured room with significant light mitigation, inside a custom-made dark box. CellSens Dimension with deconvolution (Olympus) and Imaris for Cell Biologists 9.2.1 (Bitplane) software are available on a separate HP Z8 workstation for data analysis. This microscope is excellent for simultaneous multi-color, deep, in situ imaging of excised or live samples, including tissues and whole animals under anesthesia.

Confocal and super-resolution microscopy:

‘RED BARON’ – Nikon Ti2E AXR NSPARC laser scanning confocal microscope.

Built on an inverted Nikon Ti2E microscope, the AXR NSPARC is an advanced, multi-purpose, inverted laser scanning confocal microscope with galvanometer up to 8Kx8k and resonance scanning up to 2kx2k, and Nikon spatial array confocal (NSPARC) based super-resolution imaging. NSPARC imaging can be performed using galvanometer or resonance scanning, and in super-resolution or confocal scanning modes. There are four laser lines and up to 4 fluorophores can be imaged simultaneously. Laser lines include: 405, 488, 561 and 640nm. Diascopic (brightfield/DIC) imaging is available through an independent detector.

The AXR uses four GaAsP PMT detectors. Viewing by eye uses a Nikon D-LEDI. The microscope uses a motorized X-Y stage, motorized Z-drive, Nikon Perfect Focus system generation 3, and a Prior Scientific Piezo stage (600mm travel). Available objective lenses include: 4x NA 0.2, 10x NA 0.45, 20x NA 0.80, 40x silicone NA 1.25, 60x, 60x water NA 1.27, 60x oil NA 1.40, 60x oil NA 1.49 TIRF and 100x oil NA 1.49 TIRF.

A TOKAI-HIT Cell Incubator with 1x35/50/60 mm dish and, multi-well slide and multi-well plate options is available. NSPARC imaging achieves resolutions typically near 100 nm in X-Y and 300 nm in Z. The microscope is software controlled with NIS Elements v6.02. This microscope is excellent for multi-dimensional imaging of fixed samples, precise optical sectioning, fast scanning, montaging, live imaging and super-resolution. 

‘THOR’ - Nikon TiE A1/N-SIM with TIRF.

Built on an inverted Nikon TiE microscope, the A1/NSIM is a multi-purpose, inverted laser scanning confocal microscope, TIRF and super-resolution imaging platform. This microscope uses a high-output LUNV6 laser launch. There are six laser lines and up to 4 fluorophores can be imaged nearly simultaneously. Laser lines include: 405, 445, 488, 515, 561 and 647nm.

The A1 confocal uses four PMT detectors, two conventional and 2 high-sensitivity GaAsP, while TIRF or SIM use an ANDOR-897 EMCCD detector. Widefield viewing by eye uses a SOLA Light Engine (Lumencor). The microscope uses a motorized X-Y stage, motorized Z-drive, Nikon Perfect Focus system generation 3, and a MCL Piezo Z-stage. Two-dimensional and three-dimensional structured illumination microscopy is available at 60x water immersion and 100X oil immersion. Multi-channel TIRF is available with motorized TIRF angle switching; this is not simultaneous multi- channel TIRF. TIRF is available at 100X. Available objective lenses include: 4x NA 0.2, 10x NA 0.45, 20x NA 0.75, 40x ELWD NA 0.60, 40x NA 0.95, 60x water NA 1.27, 60x oil NA 1.40, and 100x oil NA 1.49 TIRF; DIC is also possible with laser scanning microscopy.

A TOKAI-HIT Cell Incubator with 1x35 mm dish and multi-well slide options is available. Two- and three-dimensional super-resolution structured illumination microscopy (SR-SIM) achieves resolutions typically near 100 nm in X-Y and 300 nm in Z. The microscope is software controlled with NIS Elements v5.21.03. This microscope is excellent for fixed samples, precise optical sectioning, montaging small areas, super resolution imaging, and TIRF.

‘CHRONOS’ – Yokogawa CV1000 inverted spinning disk confocal microscope.

The Yokogawa CV1000 is a high-performance spinning disk confocal microscope with an enclosed, light-proof, environmentally controlled box. This microscope has 3 laser lines and is capable of outstanding brightfield microscopy via LED transmission. Laser lines include: 405, 488, and 561 nm. This microscope is equipped with a Hamamatsu high-resolution mode 1024x1024 EMCCD detector and dual Nipkow disks with 50 or 25- micrometer lens pinhole arrays that can be removed from the light path for widefield imaging. The motorized X, Y, Z stage coupled with Olympus’s Zero Drift auto focus system and software focus are great for live cell and high-temporal resolution imaging. Available objective lenses include: 4x NA 0.13, 10x NA 0.40, 20x NA 0.75, 20x LWD NA 0.45, 40x LWD NA 0.60, 40x oil NA 1.30, 60x water NA 1.20, 60x oil NA 1.35, and 100x oil NA 1.40. Three environmentally controlled and atmospheric regulated incubated chambers are available. Chambers include: 1) 1x35mm dish, 2) 3x35mm dishes and 3) low-profile multi-well plates. The microscope is controlled by Yokogawa CV1000 software and is excellent for fixed samples, dim samples, fast dynamic live samples, live tissues, organoids, embryos, multipoint montaging, and low-to-medium throughput screening in 96 or 384 well plate formats.

‘BLACK WIDOW’ - Nikon TiE inverted spinning disk confocal microscope.

This spinning disk confocal microscope is built on an inverted Nikon TiE microscope and is configured with 4 laser lines, which can be used to image up to 4 fluorophores nearly simultaneously. The laser lines are: 405, 488, 561, and 640nm. The microscope has a stand-alone 25 mW OBIS 405 nm photobleaching/photoactivation laser. This microscope uses an ANDOR-897 512x512 EMCCD detector for confocal imaging, and a Hamamatsu 512x512 EMCCD detector is available for widefield imaging. The confocal spinning disk is a Nipkow disk with an array of 50-micrometer microlenses from Yokogawa (CSU-10). A PI Nano Piezo Z-stage is available for rapid three-dimensional imaging. Viewing by eye and widefield uses a SOLA Light Engine (Lumencor).

The microscope is also equipped with a motorized X, Y, Z stage, MCL Piezo Z-stage insert, Nikon Perfect Focus system generation 3 and the following objective lenses are available: 4X NA 0.15, 10x NA 0.45, 20x NA 0.75, 40x NA 0.95, 60x oil NA 1.40, and 100x oil NA 1.45. A TOKAI-HIT Cell Incubator with 1x35 mm dish and multi-well slide option are available. The microscope is software controlled with NIS Elements v4.3 software and is excellent for fixed samples, low light imaging, fast dynamic imaging of live samples, single point photo-bleaching or -activating (405 nm laser), triggered acquisition and multipoint montaging.

Widefield, brightfield and stereo microscopy:

‘DANTE 2s’ - Olympus IX81 inverted widefield fluorescenct microscope.

This Olympus IX81 is an inverted widefield epifluorescence microscope. This microscope uses a high-intensity Excelitas XYLIS LED. Fluorescent imaging is selected using the motorized filter turrent and motorized Prior Scientific excitation and emission filter wheels, with Chroma and Semrock optical filters. The optical filters are suited for imaging offf  QD360, DAPI/Hoechst, ECFP, EGFP/Alexa488, YFP/Venus, Alexa555/568/594;TxRed;mCherry/TdTomato, Cy5/Alexa647, Cy7/Aleca740, CFP:YFP FRET, other related fluorescent molecules and some custom configurations. The microscope also has excellent brightfield and phase-contrast for the 20x and 40x objective lenses. The detector is a Hamamatsu ORCA R2 CCD and is equipped with a Prior Scientific H117 motorized, linear-encoded stage (X-Y), Olympus motorized Z-drive, software focusing, multipoint/tiling and the following objective lenses are available: 4x NA 0.13, 10x NA 0.40, 20x NA 0.70, 40x NA 0.75, 60x water NA 1.20, 60x oil NA 1.35, and 100x oil NA 1.40. For live cell imaging, an InVivo Scientific stage-top incubation chamber is available in single 35 mm dishes or multiwall plate formats, using pre-blended 5% carbon dioxide/atmosphere gas. The microscope is controlled with cellSens 4.1 software and is excellent for fixed samples and long-term timelapse imaging of live cells.

‘PEPPER’ - Nikon TiU inverted widefield fluorescent and RGB microscope.

This Nikon Ti-U is an inverted widefield epifluorescence microscope with both fluorescence and color imaging (histology). For fluorescence, this microscope uses an Excelitas Excite 100-watt metal halide lamp, Chroma optical filters, and is capable of imaging DAPI/Hoechst, EGFP/Alexa 488, TxRed/mCherry/Alexa 568/594, Cy5/Alexa 647 and related fluorescent molecules. Brightfield imaging is available, including DIC microscopy at 60X. This microscope uses a CoolSNAP HQ2 CCD detector for fluorescence. Brightfield-color microscopy is available via a Nikon DS-Fi3 24 bit, high-definition color camera (5.9 megapixel, CMOS). Other features include an ASI motorized X, Y, Z stage and the following objective lenses are available: 4X NA 0.15, 10x NA 0.45, 20X NA 0.50, 40X NA 0.75, 40x NA 0.95, 60x oil NA1.40, and 100x oil NA 1.30. This microscope is excellent for imaging fixed samples, yeast, and other cells.

'MACRO’ – Olympus MVX10 macrozoom fluorescence and RGB microscope.

The MVX10 is a flexible, high-performance macrozoom microscope equipped with fluorescence and RGB color imaging. Light source is a pE300 White with 400, 488 and 561 (Cool LED) independently triggerable LEDs and optical filters for imaging violet, green and red fluorescence emission (Chroma) via a Hamamatsu ORCA Spark CMOS camera. Brightfield-RGB imaging is via an Olympus DP23 24 bit, high-definition color camera (6.4MP, CMOS). Illuminated base with high-contrast and oblique modes. Reflected light imaging available via Fostec fiber optics. Stereo mode provides enhanced depth-of-field. Lenses include 0.63x NA 0.15, 1.0x NA 0.25 and 2x NA 0.50. Zoom body 0.63-6.3x. 2X total system magnifier. Total system magnification is 4-250x.

‘FRANK’ – Zeiss Fluorescent Stereo Microscope.

This Zeiss Discovery.V12 is a stereo microscope with fluorescent imaging. For fluorescence, this microscope uses an Excelitas Excite Series 120 metal halide lamp, and is capable of imaging EGFP, and TxRed/mCherry. There is also a FOSTEC gooseneck, halogen white light source for specific illumination. A Nikon Digital Site monochrome camera is available for imaging. Total system magnification is variable, from approximately 1.4-80X.

‘SNOOPY’ – Nikon SMZ745 zoom stereo.

The Nikon SMZ745 is a basic zoom stereo microscope with 0.67-5x zoom mechanism and an illuminated base. Total system magnification is 6.7-50x.

Analysis workstations:

‘ELEMENTS’ – Hewlett Packard Z4 v7.

Features Intel Xeon 4.6G processor-20 core, 256GB RAM, NVIDIA RTX A4000, `6GB GPU, 1TB M.2 SSD OS drive, 12TB SATA data drive. Includes Nikon Elements AR image processing, measurement analysis engine, 3D analysis, spectral deconvolution, 2D/3D deconvolution, more. 

‘IMARIS-CELLSENS’ - Hewlett Packard Z8 G4. This analysis PC features Intel Xenon 28 core processors, a 512GB SSD, 4TB HDD N 1 P4317Q data drive, 128GB DDR4 RAM, high-speed USB3.0 ports, and a Dell 43 inch Ultra HD 4K monitor. Software includes, Imaris 9.2.1 for Cell Biologists, Olympus cellSens with 3D deconvolution, FIJI, and Yokogawa CV1000 viewer.

‘HULK’ – Dell Precision 7910. This analysis PC features Intel Xenon E5-2620 6 core processors, a 512GB Samsung 850 EVO SSD, 2TB HDD, and 8TB SeaGate IronWolf data drives, 32GB RAM, high- speed USB3.0 ports, and dual Dell 24 inch UltraSharp monitors. Software includes, NIS-Elements with SIM reconstruction, MATLAB, Zeiss Zen Lite, Cell Profiler, FIJI, ICY, and Yokogawa CV1000 viewer.

The University of Colorado Boulder Library Search Engine is great for locating primary literature and text books for research. 

These short courses by iBiology Microscopy are designed for beginners.

Microforum has a wealth of information and is recommended for more advanced users. (Our very own Director is a moderator there!)

Here are some helpful microscopy-related links:

 

If you have not stumbled across the University Software Downloads and Licensing site, I highly recommend checking out the software that is available for students and staff.

Either way - here are a few well-known image viewers and advanced computer languages for image processing and analysis:

While working in the LMCF please follow the Standard Operating Procedures provided below.

From his research career, the Director has an extensive library of cell lines, fluorescent cell lines, and plasmids encoding different fluorescent reporters.

Please contact the Director at James.Orth@Colorado.edu