PAPER: PIE-scope, integrated cryo-correlative light and FIB/SEM microscopy
DEVELOPMENT OF NEW ELECTRON MICROSCOPY TECHNOLOGY INCREASES IMAGING EFFECTIVENESS, SAFETY AND ACCESSIBILITY
Understanding the biomolecules inside a cell – the way that they interact, how they organise themselves and how they give rise to life – is one of the fundamental goals of cell biology. It can help us understand diseases and disorders and help develop more effective treatments.
One major obstacle for researchers is being able to clearly see macromolecular complexes (groups of biomolecules working together as one structure). Cells are highly active, with lots of moving parts, and in order to focus on a specific region, the cell first has to be stopped via cryofreezing. A cell might also need to be thinned so the electron microscope can penetrate its layers and see the contents within. To overcome these issues scientists will often use electron and light microscopy in correlation to select, image and thin the cellular region of interest.
“It was a labour-intensive process, which required researchers to move the samples across various pieces of equipment and microscopes while keeping the sample at cryo temperatures at all times to avoid the sample being ruined,” said Dr Sergery Gorelick from Monash University’s Biomedicine Discovery Institute and lead author on the paper. “There was too much room for human error, and this is what we wanted to improve.”
Now, scientists from the Australian Research Council (ARC) Imaging Centre have designed an innovative method, which allows all critical steps to be undertaken in one place, decreasing the chances of contamination, and increasing accuracy by enabling precision thinning and instant confirmation that the area of interest hasn’t been lost in the thinning process.
The new design is an integrated cryo-Focused Ion Beam and light microscope setup called the Photon Ion Electron microscope (PIE-scope) that enables direct and rapid isolation of cellular regions.
“Up to now, essentially what we were doing was cutting and imaging blindly.” Monash University author Associate Professor Alex de Marco commented. “The microscope is something that everybody who’s ever done this type of work has been waiting for.”
“This new approach will make cryo-correlative workflow safer and more accessible,” said Professor James Whisstock, Director of the ARC Imaging Centre. “We designed PIE-scope to enable retrofitting of existing microscopes, which will increase the throughput and accuracy on projects requiring correlative microscopy to target protein complexes.”
The paper, published this month in eLife Sciences, presents the novel microscope design and software, which allows the targeting of a region in a cryo-preserved cell with a simple workflow and a precision of ~100 nanometers. The PIE-scope is creating much excitement in the field of cell and structural biology, it was downloaded by more than 500 people in the first 24 hours of publication and two companies are looking at commercialising it.
The work was a collaboration of ARC Centre of Excellence in Advanced Molecular Imaging, Monash University’s Biomedicine Discovery Institute, Ramaciotti Centre for Cryo-Electron Microscopy, Monash University’s School of Biological Sciences, Monash University’s Department of Anatomy and Developmental Biology, Biomedicine Discovery Institute, University of Warwick, EMBL Australia.
About the ARC Centre of Excellence in Advanced Molecular Imaging
The $39 million ARC-funded Imaging CoE develops and uses innovative imaging technologies to visualise the molecular interactions that underpin the immune system. Featuring an internationally renowned team of lead scientists across five major Australian Universities and academic and commercial partners globally, the Centre uses a truly multi-scale and programmatic approach to imaging to deliver maximum impact.
The Imaging CoE is headquartered at Monash University with four collaborating organisations – La Trobe University, the University of Melbourne, University of New South Wales and the University of Queensland.
About the Monash Biomedicine Discovery Institute
Committed to making the discoveries that will relieve the future burden of disease, the newly established Monash Biomedicine Discovery Institute at Monash University brings together more than 120 internationally-renowned research teams. Our researchers are supported by world-class technology and infrastructure, and partner with industry, clinicians and researchers internationally to enhance lives through discovery.
Written by Aimee Sanderson
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