One of the problems associated with imaging fluorescence in large biological samples is the obscuring effects of light scatter. Traditionally this has meant physically sectioning the material into optically-thin slices in order to visualise microscopic structure. With the advent of new volumetric imaging techniques, e.g. lightsheet microscopy, there is increasing demand for procedures that allow deeper interrogation of biological tissues. With this in mind, an innovative clearing system has recently been purchased through generous donations to the European Cancer Stem Cell Research Institute (ECSCRI). The equipment, which will be housed in ECSCRI lab space, allows large, intact histological samples to be rendered transparent for fluorescent labelling and 3D visualisation by confocal and lightsheet microscopy.
The X-Clarity tissue clearing system is designed to simplify, standardise and accelerate tissue clearing using the CLARITY technique (an acronymn for Clear Lipid-exchanged Acrylamide-hydridized Rigid Imaging/Immunostaining/in situ-hybridization-compatible Tissue hYdrogel). In the technique, preserved tissues are first embedded in a hydrogel support matrix. The lipids are then extracted via electrophoresis to create a stable, optically transparent tissue-hydrogel hybrid that permits immunofluorescent labelling and downstream 3D imaging.
The new equipment and associated reagents will have wide relevance to many areas of research in Cardiff, including deep visualisation of breast cancer tumours by Professor Matt Smalley’s research group using the Bioimaging Hub’s new lightsheet system. You can see a video here that shows the power of the CLARITY technique for high resolution 3Dvisualisation of tissue and organ structure.
Above: The new Ultimaker ‘Extended’ 3D printer building a stage insert for a novel microscope system.
The Bioimaging Research Hub has recently purchased an Ultimaker 3 ‘Extended’ professional 3D printer. The printer will allow users to 3D print bespoke pieces of scientific equipment or generate scale models of microscopic samples for use in their research as well as for teaching, outreach and engagement activities (examples can be found here andhere).
The 3D printer utilises FDM (Fused Deposition Modelling) printing technology and has a range of advanced features allowing the fabrication of professional quality, high resolution 3D prints. The printer can print in two different colours or a single colour in addition to a dissolvable PVA support scaffold, thus allowing complex overhanging structures to be printed at high fidelity whilst significantly reducing finishing time.
The printer has a large build volume (215 x 215 x 300mm), supports a range of materials (nylon, PLA, ABS, CPE and PVA) and has a print resolution of 20-200 microns. The printer is wi-fi enabled with an internal webcam so that users can remotely monitor the progress of their 3D prints.
Above: ‘Cutting-edge’ equipment: the Zeiss PALM MicroBeam laser micro-dissector located at the European Cancer Stem Cell Research Institute (ECSCRI)
A Zeiss PALM MicroBeam laser micro-dissector system is now available as a spoke of the Bioimaging Research Hub. The equipment, which is located at the European Cancer Stem Cell Research Institute (ECSCRI), allows isolation of DNA, RNA and protein from laser micro-dissected samples from both histological sections (paraffin wax or cryo) and live cells. Further details of the system are available through our equipment database. All enquiries for this system should be directed towards Mr Mark Bishop.
The old electron microscopy darkroom (BIOSI 2; E/0.05) within the Bioimaging Research Hub has recently been refurbished as a live cell imaging suite via generous support from BIOSI. It now houses a spinning disc confocal system for fast, live cell imaging applications. The system is based around an Olympus IX71 inverted microscope, kindly provided by Dr Pete Watson, which has been upgraded, via ISSF funding, with a Crest Optics X-Light V2 confocal head, a Cairn Research tri-line laser bank (405nm, 488nm, 561nm) and a Hamamatsu ORCA Flash 4 sCMOS digital camera with M-View Gemini image splitter. The system is fully integrated via Molecular Devices MetaMorph software and boasts a 40″ 4k display. The system will expand the Hub’s imaging toolbox, enabling high speed, multi-position, multi-colour 3D/4D image acquisition. Support systems for live cell imaging (i.e. gas and incubation) are also available within the facility. Further details of this system are available through our equipment database.
Imagine being able to generate a highly accurate, solid scale replica of the sample that you are visualising down the microscope; a perfectly-rendered pollen grain, or blood cell, or microscopic organism, but big enough to hold and examine in your hand. It would allow much better 3D conceptualisation of the sample, particularly for blind or visually-impaired individuals, and would have enormous utility in teaching and in engagement activities, and what researcher wouldn’t want a tangible, physical embodiment of their research to help explain their work (and impress their colleagues) at scientific meetings? Sounds like the stuff of science fiction doesn’t it? Well, not any more. Thanks to 3D printing technology (and the help of Dr Simon Scofield‘s lab) we have started taking volume datasets from the confocal microscope out of the virtual world and making them a reality. If you would be interested in generating a highly accurate scale model of your favourite biological sample (or would simply like to handle a giant pollen grain!) then please feel free to get in touch.
Dr Amit Jathoul and Professor Jim Murray (BIOSI) have recently obtained generous funding from the Wellcome Trust to purchase a real-time, multi-spectral in vivo imaging system from Biospace Lab. The PhotonIMAGER allows bioluminescence/fluorescence imaging at a wide range of scales from cells, tissues and organs to entire complex organisms, thereby bridging the gap from single cell to whole organism imaging. Further details of the system are available through the research equipment database. All enquiries for this system should be directed towards Dr Amit Jathoul.
The BIOSI Bioimaging facility has recently expanded its imaging toolbox with a new, state-of-the-art confocal microscope system, that was purchased through generous funding by the Research Infrastructure Fund (Lead Applicant: Dr Walter Dewitte). The system, a top-of-the-range Zeiss LSM 880 upright confocal microscope, features the advanced Airyscan super-resolution detection module which provides a 1.7x gain in resolution in all three dimensions compared to conventional confocal optics. The system also supports advanced fluorescence techniques including FCS (fluorescence correlation microscopy) and FLIM (fluorescence lifetime imaging (FLIM) – the FLIM module will be installed during the first week of December. Further details here.