The Centre for Microscopy, Characterisation and Analysis (CMCA) is a core facility centre within The University of Western Australia, where research excellence is achieved by providing world-class science infrastructure and expertise to researchers and industry. Provision of such excellence is achieved through a focus on world-class facilities matched with expertise, and through a concept-to-publication User Pathway designed to meet the needs of researchers and industry through quality in training, acquisition and analysis.

The Centre comprises 35 academic, research, technical and administrative staff situated across four sites and supporting more than 45 instrument platforms. These platforms include secondary ion mass spectrometry (SIMS), optical and electron microscopy, cytometry & cell sorting, small animal imaging, micro-CT and MRI, confocal Raman microscopy, NMR, powder, thin film and single-crystal X-ray diffraction, bio-organic mass spectrometry, and scanning probe microscopy.

Located within the Harry Perkins Institute of Medical Research in Nedlands, the [email protected]EII node houses the light microscopy infrastructure (as well as the flow cytometry, small animal imaging and MRI facilities), including the newly installed super resolution microscopy suite, featuring three high-end platforms. The location is nestled within a medical research precinct, making these platforms easily accessible by researchers from all of the universities and research institutes in Western Australia and, more widely, Australia.

Nikon A1Si spectral detector confocal system plus TIRF on a Nikon Ti-E inverted motorised microscope.

This inverted confocal microscope is equipped with 4 solid state lasers (405nm, 488nm, 561nm and 640nm), four photomultiplier fluorescence detectors, a transmission detector (can perform DIC, polarization or phase contrast imaging), as well as a 32-channel spectral detector. Imaging can be performed either in simultaneous or sequential mode, and we have a combination of high quality dry and immersion objective to suit your imaging needs. Complex imaging techniques are able to be performed on this system, including multiple fields, montaging, reflection and Fluorescent Recovery After Photo-bleaching (FRAP) microscopy.

Long term time-lapse imaging is possible with the use of the Tokai Hit stage top incubator in combination with the Perfect Focus System (PFS) to prevent focus drift. Two colour (488nm and 561nm) total internal reflection fluorescence (TIRF) microscopy is integrated into the microscope and high-speed image capture is available using the Andor 885 EMCCD camera, all of which is controlled by the NIS Elements software.

In addition to this, we also have a Nikon DS-2Mv colour camera which can be mounted onto the frame, and used for static or live cell imaging.

Nikon A1Si spectral detector confocal system

NikonA1 RMP on Ti-E (inverted)

Nikon A1 RMP on Ti-EThis inverted system is a dual system, capable of both confocal and multiphoton imaging. It comprises of four solid state lasers and four photomultiplier fluorescence detectors, as well as a Coherent Chameleon II pulsed laser (700-1000nm), and four NDD detectors. Furthermore, there are both galvanometer and resonant scanners on this platform to facilitate various imaging speed requirements.

To perform in vivo animal imaging, we have an objective inverter, with an accompanying external stage. Dipping lenses of various magnifications are also available, including the Nikon Apo 25x water immersion multiphoton objective.

Nikon A1R with PicoQuant FCS/FLIM in Ti-E (inverted)

This inverted confocal has Picoquant module attached to enable Fluorescent Lifetime Imaging (FLIM) and Fluorescence Correlation Spectroscopy (FCS) imaging. FLIM is a technique which investigates the exponential decay rate, or lifetime, of a fluorophore, which is sensitive to changes in the local environment. FLIM is also used to interrogate FRET pairs, providing a stronger correlation than analysis by confocal means. FCS enables the investigation of single molecule diffusion dymanics within a femtolitre volume.

This system comprises of four solid state lasers with two photomultiplier fluorescence detectors and two Galllium Arsenide Phosphatide (GaAsP) detectors for confocal imaging, as well as three picosecond pulsed lasers (405nm, 485nm and 640nm), with two PMA hybrid detectors. Furthermore, there are both galvanometer and resonant scanners on this platform to facilitate various imaging speed requirements. FLIM and FCS images are acquired using PicoQuant Symphotime.

Live cell imaging can also be accommodated on this platform via the use of an Oko Labs stage top incubation system, and Perfect Focus System (PFS)

Techniques

  • Four colour confocal microscopy
  • Fluorescent Lifetime Imaging (FLIM)
  • Fluorescence Correlation Spectroscopy (FCS)
  • Multipoint and live time-lapse imaging
  • High speed resonant scanning
  • JOBS multidimensional acquisition software

Instrument specifications

  • Inverted Nikon TiE microscope
  • Nikon PlanApo objectives 10x-40x, air and oil immersion
  • Nikon PlanApo 60x WI IR, 1.27NA objective
  • Nikon PlanApo 100x Oil immersion, 1.45NA objective
  • Four 50mW solid state lasers – 405nm (violet), 488nm (blue), 561 (yellow), and 640nm (red)
  • Two PMT detectors
  • Two Gallium Arsenide Phosphatide (GaAsP) detectors
  • Galvanometer and resonant scanners
  • Three picosecond pulsed lasers – 405nm (violet), 485nm (blue) and 640nm (red)
  • Picoquant Symphotime 64 analysis software
  • Two PMA hybrid detectors
  • Piezo stage
  • Perfect Focus System III
  • NIS Elements software controlling confocal acquisition and lasers
  • Oko Labs stage top incubator
Nikon A1R with PicoQuant

Nikon SIM (Ti2) (inverted)

Nikon SIM Ti2Housed on a state-of-the-art Ti2 microscope, our SIM platform can acquire images with a resolution down to 100nm. Various objectives are available for this system, including 100x oil immersion with motorized correction collar, and 60x WI objective, specialized for live cell imaging. It is a four colour platform (405nm, 488nm, 561nm and 640nm solid state lasers), where images are acquired on an Andor iXon 897 EMCCD camera.

In addition to this, the system is capable of imaging in either 2D SIM, 3D SIM, 3D stack SIM or TIRF SIM mode for fixed or live cells (Tokai Hit stage top incubation system). Stability of the field of view over the imaging period is enabled by the Piezo stage and Perfect Focus System (PFS).

Techniques

  • 2D SIM
  • 3D SIM
  • 3D TIRF SIM
  • 3D SIM Stack
  • Live cell imaging

Instrument specifications

  • Inverted Nikon Ti2 microscope
  • Nikon PlanApo objectives 10x-20x, air
  • Nikon SR PlanApo IR 60x 1.27NA WI objective
  • Nikon SR Apo TIRF 100x 1.49NA Oil immersion objective
  • Four solid state lasers – 405nm (violet, 67mW), 488nm (blue, 120mW), 561 (yellow, 120mW), and 640nm (red, 80mW)
  • Andor iXon897 EMCCD camera
  • Three & four colour grating blocks for 60x WI & 100x Oil objectives
  • Piezo stage
  • NIS Elements software controlling confocal acquisition and lasers
  • Tokai Hit stage top incubator

Nikon STORM (inverted)

Nikon STORM Localisation imaging is the final super resolution microscope platform in our suite. Imaging with this modality exploits the photophysical behaviour of fluorophores to Our N-STORM system is built on an inverted Ti-E microscope, fitted with four solid state lasers (405nm, 488nm, 561nm and 640nm) and a Hamamatsu OCRA FLASH 4 sCMOS camera, for high speed acquisition. C-STORM imaging can be performed on this platform, as well as the novel N-STORM imaging modality, where tandem dye pairs are used conjugated to samples utilising the high performance of Alexafluor 647 fluorescent dye.

The system is also equipped with Tokai Hit stage top incubator for live cell imaging, and Perfect Focus System (PFS) to prevent drift.

Techniques

  • C-STORM
  • N-STORM
  • STORM with TIRF
  • Live cell imaging

Instrument specifications

  • Inverted Nikon TiE microscope
  • Nikon PlanApo objectives 10x-20x, air
  • Nikon HP Apo TIRF 100x 1.49NA Oil immersion objective
  • Four solid state lasers – 405nm (violet, 67mW), 488nm (blue, 120mW), 561 (yellow, 120mW), and 647nm (red, 300mW)
  • Hamamatsu OCRA FLASH 4 sCMOS camera
  • Piezo stage
  • Perfect Focus System III
  • NIS Elements software controlling confocal acquisition and lasers
  • Tokai Hit stage top incubator

Nikon Ti (inverted) with 6-colour fluorescence

This epifluorescence platform is utilising an inverted Ti-E microscope, with a special combination of filters and dichroics to be able to rapidly image six fluorescent probes. Using a Prior encoded motorised stage and an Andor iXon 885 EMCCD camera, whole section, six-coloured fluorescence imaging and analysis can be performed.

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