Light and laser microscopy facilities - The University of Sydney
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Infrastructure_

Light and laser microscopy

Capabilities in optical microscopy
Access an extensive range of light and laser microscopes across the campus to meet your research objectives.

At Charles Perkins Centre

THUNDER Imager features Leica technology Computational Clearing. It efficiently removes out-of-focus blur in real time, enabling the meaningful interrogation of 3D specimens with camera-based wide-field fluorescence microscopes.

Microscope body:
DMi8 Inverted microscope with Quantum HS linear motorised stage. THUNDER closed-loop Auto. Focus Control.

Source of light:
LED 8.
395 nm, 438 nm, 475 nm, 511 nm, 555 nm, 575 nm, 635 nm, 730 nm
DAPI, FITC, TRITC, Cy5, mCherry, CFP, YFP, Cy7

Cameras:
Leica K8 back-thinned sCMOS camera. 2048 px x 2048 px, 6.5 ÎĽm pixels. 95% QE.

Objectives:
5X/0.12, 10x/0.32, 20X/0.80, 20X/0.40 CORR, 40X/1.010 W CORR, 63X/1.40 Oil

Carriers:
Multi-well plates, chamber slides, glass slides, petri dishes.

Contrast methods:
DIC, Phase

Controlling software:
Leica LASX with Navigator & 3D visualisation modules. 

A Zeiss upright fluorescent microscope Axiolmmager.Z2 integrated with MetaSystems SlideFeeder X80, Metafer software with VSlide, deep neural networks (DNNs) Classification and DNN Server software licences. It comes with automated bar code reader and slide label camera.

Using Metafer software, high-throughput images can be automatically captured using brightfield or fluorescent modalities. This system is a highly specialised unit capable of imaging and analysing data sets with the DNN with maximal efficiency.

The MetaSystems currently has 256 slides capability (160 standard and 96 custom-built), calibrated with slide feeder robot. This slide scanner is equipped with solid state self-calibrating and stabilised LED fluorescence light source with 7 excitation lines: 385nm, 430nm,  475nm, 555nm, 590nm, 630nm, 735nm. High resolution objectives include: 5x/0.25, 10x/0.45, 20x/0.8 and 40x/1.3. Oil, with automated immersion oil dispenser.

This instrument is also equipped with MetaSystems CoolCube 4m monochrome camera, captured 12-bit images with a size of 4,096 x 3,000 pixels. The CoolCube 4 camera models are ultra high-resolution digital CMOS cameras using a 1.1" CMOS chip with a pixel size of only 3.45µm x  3.45µm.

It features Spinning Disk (SD) Confocal with Super-resolution by optical ReAssignment (SoRA), Total Internal Reflection Fluorescence microscopy (TIRF), Adaptive optics (AO) and Phasor holography for photomanipulation applications. The Laser launch, AO, holography, rapid switching (~1 ms) of light path and complete software control on the system are fully integrated.

Microscope body:
Nikon Ti2-E Inverted with Perfect Focus 4. Motorised X,Y stage. Piezo Z stage (300 ÎĽm range)

Full blackout enclosure incubator with temperature, CO2 and humidity control. DIC capable

Laser lines:
405 nm, 488 nm, 514 nm, 560 nm, 594 nm & 642 nm

Cameras:
SD confocal module features four Photometrics Prime 95B back illuminated sCMOS cameras for up to four channels simultaneous image acquisition. 

TIRF modules is equipped with two Photometrics Prime 95B back illuminated sCMOS cameras for the simultaneous imaging of two different cellular markers.

Nikon Objectives:
10x/0.45, 20x/0.75, 25XC/1.05 Silicon, 40XC/1.25 Silicon, 100XC/1.35 Silicon
60x/1.49 Oil TIRF, 100XC/1.49 Oil TIRF

Controlling software:
SlideBook

A Microscope for High-Resolution, Fast, and Gentle 3D Live Cell Imaging. The system generates an optical lattice to create an ultra-thin light sheet to image biological samples over long periods of time and with very fine resolution.  It uses Bessel beam lattice sheet illumination via cylindrical lenses and high-speed SLM for multicolor-imaging; annular mask array for various light sheets; galvo mirrors to control lattice movement in X and Z; X,Y translation stages and piezo imaging objective control.

Laser lines:
405 nm, 488 nm, 560 nm & 642 nm

Cameras:
Two Hamamatsu ORCA Fusion BT sCMOS cameras.  2304 px x 2304 px, 6.5 ÎĽm pixels, 95% QE.

Objectives:
Illumination objective. Custom-made 28X/0.71NA water immersion objective.

Detection objective. 25X/1.1NA water immersion objective.

Sample holder:
Cells should be seeded onto 5 mm coverslips. Imaging area: 3 mm x 3 mm region from the centre of the coverslip. Volume: ~150 ÎĽm.

Okolab Incubator with Bold Line temperature controller, heater and digital CO2 gas mixer for comprehensive environmental control.

Controlling software:
SlideBook

Super Resolution Fluorescence Microscopy collects images of fluorescent molecules with resolution that is not limited by the diffraction limit of light. Stimulated Emission Depletion (STED) is built on a Confocal microscope and is a laser scanning technique.

STED can improve lateral resolution to a theoretical limit of 50nm, actual ~100nm. Axial resolution can also be improved with STED to a theoretical limit of 130nm, actual ~300nm. If optimal resolution in XY and Z is required the actual achievable resolution is approximately 150nm laterally and 500nm axiallly.

This microscope also features an Insight DeepSee laser for multiphoton and Second Harmonic Generation (SHG) imaging with excitation wavelenghts of 680-1300nm.

Black paneled box incubator around microscope for live cell imaging.

Lasers
  • UV: 405nm 
  • Argon: 458, 476, 488, 496, 514nm
  • White Light Laser (WLL): tunable 470nm - 670nm
  • Multi-photon 1: tunable 690nm - 1300nm
  • Multi-photon 2: 1040nm
  • STED 592nm
  • STED 775nm
Detectors
  • Internal, AOBS: tunable 400nm - 800nm
    • 3 standard PMTs
    • 2 Hybrid Detectors (high sensitivity PMTs)
  • External Non-Descanned (uses filter cubes)
    • Transmitted Light: 2 standard PMTs
    • Reflected Light: 2 standard PMTs & 2 Hybrid Detectors

Confocal lasers: 403, 457, 488, 514, 560, 640nm

FLIM / FCS / FCCS lasers: 405, 488, 640nm

Detectors

DU4 uses Filter Blocks

  • 2 standard PMTs
  • 2 Galium Arsenide Phosphide (GaAsP; high sensitivity PMTs)

Spectral Detector

  • uses prism to split emission wavelengths to discrete bandwidths
  • 2 FLIM / FCS / FCCS GaAsPs

Total Internal Reflection microscopy is a Super Resolution technique.

Specifications
Lasers 405, 488, 561, 640nm
Lumencor SpectraX LED 395, 440, 470, 508, 555, 640nm
DMD for photoactivation and photouncaging
Cameras TIRF Monochrome - Andor iXon Ultra 888 (max 26 f/s at 1024x1024, 96f/s at 512x512)

Visualise, analyse, quantify and phenotype immune cells in situ.

The Mantra quantitiative pathology workstation, with inForm image analysis software, enables easy visualisation, quantification and phenotyping of cells in situ in tissue sections using multiplexed biomarkers. This integrated workstation features a spectral detector to collect spectral data for unmixing overlapping signals and removal of autofluorescence. inForm software features user-trained algorithm to enable automatic identification of specific tissue types based on tissue morphology and indentification of cells within the tissue for quantification.

Upright Confocal microscope with motorised stage.

Lasers: 405, 488, 561, 640nm

Detectors: 3 standard PMTs using Filter Blocks - DAPI/Cy5, FITC, TRITC

Upright microscope for mosaics and multipoint.

Light Source: Lumencor SpectraX LED - 395, 440, 470, 508, 555, 640nm

Cameras: Monochrome - DS-Qi2, Colour - DS-Fi2

Incucyte SX5 can automatically acquire and analyse in a variety of formats within the stable environment of a tissue culture incubator. It supports 3 interchangeable vessel trays and over 700 vessels, compatible with flask, dishes, slides and microplates - up to 6 microplates in parralel.

It can support up to 5 different fluorescence channels, up to 3 at a time (Green/Orange/Near-IR) plus HD Phase imaging for long term time-lapse experiments. There is also the option of a Green/Red Optical Module.

The Incucyte SX5 also has a turret for the 4X/10X/20X objectives, as well as supporting independent scheduling of plates. Multi-user, multi-application support through remote network capability, unlimited free user licences, independent vessel scheduling and multi-objective scheduling.

Additional Software Modules:
1. Incucyte® Cell-by-Cell Software Module
2. Incucyte® Advanced Classification Module
3. Incucyte® Spheroid Software Module
4. Incucyte® Angiogenesis Software Module
5. Incucyte® Scratch Wound Software Module including
    Incucyte® Cell Migration Kit

This automated slide scanner microscope can scan up to 100 slides at one time with brightfield or fluorescence mode. The system is deisgned specifically to digitise microscope slides for your research. It can also perform a Z-stack of up to 50um, to achieve a fully stitched 3D construction of the tissue of interest.

Filterwheel  Excitiation (nm)   Beam Splitter (nm)   Em (nm)
DAPI                G365                      FT 395                         445/50
GFP                 BP 470/40             FT 495                         525/50
DsRed             BP 550/25             FT 570                         605/70
Cy5                  BP 640/30            FT 660                         695/50
 

 

Live cell fluorescence and confocal laser scanning microscope with Airyscan detector for super resolution. Has a live cell incubator cage on the microscope to keep cells at appropriate temperature and CO2.


Super resolution: Lasers 405,488,561, 640nm and tuneable emission range 400-700nm with Airyscan detector for super resolution down to ~125nm XY and ~350nm Z.

Laser scanning and confocal microscopy: in addition to the Airyscan GaAsP PMT detector which can be used as a standard detector, there are two GaAsP PMT and a transmission detector with DIC optics. The system has tuneable emission wavelength selection between 400-700nm with clean up emission filters available.

Spectral Imaging and Unmixing: The system is capable of spectral imaging emissions from 400nm-700nm and unmixing for 5-6 channels. This works well for one blue, three green and red and one far red probe as well as autofluorescence.

Widefield fluorescence microscopy: This system has a metal halide white light source and monochrome camera for widefield fluoresence imaging.

The AZ100 covers an extremely wide range of magnifications, from 5x to 400x. This microscope can continuously switch magnifications, extending from macro to micro observation of the same specimen.

Widefield and Spinning Disk confocal live cell fluorescence, DIC, large mosaics, multi-point, time series, perfect focus.

At Madsen Building

Equipped with 405,488,561 and 640nm lasers and a transmitted ligt detector allwing for the majority of fluorophores to be imaged with a beautiful DIC (or brightfield) overlay. With the fastest Resonance Scanner on the market, it can capture up to 438fps at 512 x 32 or 30fps at 512 x 512. This, combined with a stagetop incubator results in excellent temporal resolution for imaging live cells.

Live cell imaging is made easy with Multiple Area Time Lapse acquisition and Micropplate navigator. TruSpectral technology prvovides fantastic spectral resolution that enables users to collect the emission profile of a visible fluorophore at any point in their sample. It can also be used for up to 16 channel sequential scanning. This gives rise to a spectral un-mixing function which allows separation of spectrally close fluorophores, particularly handy for users with autofluoesence interfering with the signal of their labelled target of interest.

This is a conventional inverted fluorescence microscope equipped with motorised stage with definite focus. The Shuttle & Find from Zeiss is a correlative microscopy interface for light and electron microscopes. It can be used with the Zeiss Sigma HD SEM.

Inverted optical microscope for materials science. Ample working space to easily place large and heavy samples (up to 30kg). DMC4500 camera, reflected light only, DIC, Polarizer, manual objectives and stage, Leica LAS-X software.

This instrument covers a broad range of confocal and multiphoton imaging requirements. Including TauSense, a set of tools based on fluorescence lifetime information, it provides an additional contrast, improved image quality and separation of spectrally overlapping flurophores. White laser light for single photon excitation tunable from 440-790nm. Extended detection range up to 850nm.

Equipped with:
- Stellaris 8 DIVE technology, including a Spectra-Physics Insight X3 Single Ti:Sapphire femtosecond pulsed laser, specialised objectives and 4Tune spectrally tunable non-descanned detectors which enables imaging of more than 1mm microns deep into thick specimens without any signal drop-off. Multiphoton excitation variable 680-1300nm.
- Stellaris 8 FALCON technology that provides fast FLIM at confocal speed along with FLIM analysis tools, such as exponential fitting, FLIM phasor analysis, pattern fitting and FLIM-FRET analyzer. Stellaris 8 FALCON allows to quantify fast cellular processes, like molecular interaction or molecular environmental changes.

Features a tandem scanning system to provide highest axial resolution with galvanometric scanner, switchable between resonant and non-resonant mode. With the galvo scanner we can image up to 1892 x 8192 (64M pixel) array. The resonance scanner allows imaging up to 512 x 512 pixel array at 28 frames per second or 512 x 16 at 290 frames per second. This high-speed raster scanning system is particularly well suited to live samples and physiological measurements.

 

 

At Brain and Mind Centre

This custom built system is the world's first commercially available multiphoton system to contain 3 tuneable lasers. Its special lasers can peer deep into the tissue of live animals, providing researchers with a view of biological or pathological processes as they occur. It uses 3 nano-second pulsed lasers which can illuminate up to 8 fluorophores simultaneously, providing exceptionally fast imaging.

This state of the art slide scanner allows researchers to observe labelled tissues on slides, using fluorescence, Brightfield, dark field and polarisation modalities. It offers the choice to acquire immunoflourescently labelled tissues, or typical staining procedures including DAB and H&E stained tissue.

The system has a robotic arm used in conjunction with slide holders for autoloading up to 210 slides. The system autofocuses onto the tissue on the slides, recognises the edges of the tissue and scans the entire tissue, stitching the image in real time before moving onto the next slide. It can perform a Z-stack, to achieve a fully stitched 3D construction of the tissue of interest. This slide scanner provides high resolution imaging with a 60x Silicon Oil objective with auto-oiler.

This slide scanner has the capacity to scan 400 slides in one run, saving the files directly onto a special server. The system only has a 20X objecitve (with capacity for doubling to 40X) and only performs brightfield scanning. Performs slide scanner with a fully automated system and is very simple to operate.

Equipped with 405,457,488,514,640nm lasers and a transmitted light detector allowing for the majority of fluorophores to be imaged with a beautiful DIC (or brightfield) overlay. This microscope has both a stagetop and cage incubator resulting in excellent stability for imaging living cell dynamics 24,48 even 72 hrs. Live cell imaging is made easy with Multiple Area Time Lapse acquisition. Set up the microscope to image live cells from each well in a multiwell plate overnight.

We have 2 upright confocal microscopes with motorised stage ideal for imaging fluorescent slides. The motorised stage can be used to collect large stitched mosaics combined with z-stack function for very large volume images.

Basic upright widefield fluorescence microscope for mosaics using a combined monochrome/colour camera for fluorescence and histology. It uses a white light source (mercury lamp) and a set of filter cubes.

Filter Cubes      Ex                       Dichroic         Em
Ana
DAP                    BP 350/50          LP 400           BP460/50
L5                       BP 480/40          LP 505            BP 585/40
RHO                   BP 546/10           LP 560           BP 546/10
Y5                       BP 620/60          LP 660           BP 700/75

 

Fluorescence Stereomicroscope for brightfield or fluorescence with 0.63x and 2.0x objectives and 3 filter positions for blue, green and red fluorescence. Motorised zoom 16:1.

At Kolling Institute

This instrument is designed for high precision cutting of fresh and/or fixed tissue and cells. It can be used to isolate regions of interest (ROIs) for downstream genomics, transcriptomics, proteomics and metabolomics. This system can also be used with live cell culture for cloning or recultivation of single cells.

This system is equipped with brightfield or blue and green fluorescence filter cubes to find your ROIs. Can be used with a range of collector vessels including 0.2 or 0.5 ml tube caps, through 8 and 12 cap-strips up to standard 96-well and 386-well plates (max. 352 wells addressable).

Basic inverted widefield fluorescence microscope with manual stage. It is equipped with a Colour camera for brightfield and a monochrome camera for fluorescence. It uses a white light source (metal halide lamp) and a set of filter cubes including DAPI, GFP, YFP, and RFP.  Available objectives include 1.25x, 5x, 10x, 20x, 40x.

This instrument covers a broad range of requirements including brightfield, widefield and confocal fluorescence for live or fixed samples. Including TauSense, a set of tolls based on fluorescence lifetime information, it provides an additional contract, improved image quality and separation of spectrally overlapping flurophores. White light laser for single photon excitation tunable from 485-790nm. Extended detection range up to 850nm.

Features a tandem scanning system to provide highest axial resolution with galvanometric scanner, switchable between resonant and non-resonant mode. With the galvo scanner we can image up to 8192 x 8192 (64Mpixel) pixel array. The resonance scanner can image up to 512 x 512 pixel array  at 28 frames per second or 512 x 16 at 290 frames per second. This high-speed raster scanning system is particularly well suited to live samples and physiological measurements.


Enquiries

For further information on our light and laser microscopy capabilities, please contact: smm.lo@sydney.edu.au