Dr Peter O'Toole: Taking technology further
From imaging to cytometry, Dr Peter O'Toole from the UK-based Bioscience Technology Facility has taken microscopy out of the lab and into the future. [Image: Imagingscientist.com]
Dr Peter O'Toole's passion for practical science didn't start at school. The sciences weren't his favourite subjects and while excelling in some areas, he didn't, as he puts it, 'ace' chemistry.
"My brother studied Biochemistry at Imperial College London, and not knowing what to do after my A-Levels, this inspired me to do a degree in biochemistry as well," he recalls. "It sounded diverse, it sounded interesting and it didn't put me down any one route, which at the time, was good."
Come his final year at University of Essex, UK, and still unsure of his future, the young student came across spectrin, a protein largely responsible for the characteristic bioconcave shape of red blood cells.
"I didn't know where I was going with my career and was just focused on finishing my degree," he says. "But my third year research project really gave me the bug for research. I had suddenly found a real passion for science; all the learning suddenly made sense and research brought everything together - I just wanted to learn more."
Working under biochemist, Professor Richard Cherry, O'Toole's undergraduate studies into fluorescence polarisation of spectrin led to a PhD, investigating lipid interactions in the same protein, and eventually dynamic studies with fluorescence microscopy.
"Actually, I'd always avoided microscopes - I had never got them focused right, everything would be blurry and I thought they were just awful," he laughs. "But then I looked down a good microscope that was aligned correctly and a new world opened up."
"I could see how everything worked and that was brilliant," he adds. "We were doing single particle studies and I suddenly I was looking at starry skies, the tiny fluorescent spots on a slide. Although not simple to execute, I personally found it obvious to use and was immediately at one with the microscope."
Time-lapse ptychographic phase imaging: a) Brightfield, Zernike phase contrast, DIC and ptychographic phase image of neuronal cultures. (b) Brightfield images acquired at various focal positions by moving objective in the z axis. (c) Maturation of primary hippocampal neuron cultures imaged in time-lapse starting at day 1 in-vitro. [scale bar = 100 μm].
From Time-lapse ptychographic phase imaging of rat hippocampal neuronal cultures; [Suman et al, Scientific Reports volume 6, Article number: 22032 (2016)]
PhD led to postdoc and an eventual Wellcome Trust project with a difference. Provided with a confocal microscope and flow cytometer, O'Toole was able to develop new FRET and FRAP approaches in collaboration with life science research business, Bio-Rad.
"I loved spectrin but felt there was only so much I could do with one protein," he says. "I'd realised that I was really interested in the technologies and having been given a postdoc without portfolio; it was brilliant."
With instrument developed and successful beta tests in tow, backed up by an 'inspirational' advanced imaging workshop at the European Molecular Biology Laboratory, O'Toole was smitten with technology and felt he had finally found his future. Still, it was 2002, technology-oriented posts were few and far between and the concept of a 'core facility' had yet to emerge. Thankfully, for the up and coming researcher, this would soon change.
Single-particle tracking of chemokines in collagen. Second-harmonic imaging microscopy of collagen network in (A) 2D and (B) 3D. (C) Representative consecutive submillisecond images of chemokines in collagen. Adapted from High-Speed Single-Molecule Tracking of CXCL13 in the B-Follicle; [Miller et al, Front. Immunol., 22 May 2018.]
At the time, the University of York was looking for a researcher to lead the Imaging and Cytometry labs in its soon to open Bioscience Technology Facility. O'Toole applied, interviewed and was offered the job the next day.
"At the time, researchers would have, say, a confocal microscope and flow cytometer, but these were often found in individual laboratories and the research councils were not yet encouraging core facilities," points out O'Toole. "But York had a vision to build a technology hub where all the necessary staff and technology would reside in one place, and researchers wouldn't have to worry about looking after bits of kit and post-docs leaving and taking their high-end technology expertise."
"Of course this was when technology suddenly started to explode," he adds. "York was right there from the start, made it work and is now a financially sustainable facility attracting funds and great scientists."
In the lab: Peter O'Toole [Imagingscientist.com]
According to the researcher, in the beginning, the entire facility was simply an empty shell. But in the last sixteen years it has attracted more than £17 million equipment, with a hefty share funding light and electron microscopy, as well as flow cytometry, within the Imaging and Cytometry laboratory.
"Being there from the start has been the best apprenticeship you could do," says O'Toole. "At the time, the user demands were not so high and the technology was more edgy and we really had to get to grips with the equipment to fix the technical problems. Now we're just full-on and flat out with users."
Today, the entire Bioscience Technology Facility spans some 2000 m2, and as well as Imaging and Cytometry, provides laboratories in Molecular Interactions, Protein Production, Genomics and Bioinformatics, and Metabolomics and Proteomics.
"I have always wanted to be involved with technology and this job has really provided that opportunity," adds O'Toole. "In the last decade, the research councils have really responded with funds, and now you can have a career in developing technology and applications. It's still rare and isn't easy, but it is possible. York has been exceptional at providing and supporting this opportunity and provides a compelling model for other Universities to follow."
Since arriving in York, O'Toole has been involved in myriad bioimaging projects. For example, he has developed techniques using laser scanning microscopy to image and manipulate biofilms and adapted electroporation to image mammalian cells in vivo. Meanwhile, he has also been a driving force behind the development of two-photon intravital imaging for brain metastases studies at York.
But for the researcher, the real excitement has resided in two key projects; the £1.55 million Research Council-funded 'Bio-Continuum Microscopy' and label-free imaging using novel instrumentation.
The former focuses on bringing light and electron microscopy onto a single fluorescence electron microscopy platform to image from the micro- to the nano-scale. Here, O'Toole has collaborated with microscopists and researchers from The Francis Crick Institute in close collaboration with Lucy Collinson, correlative light and microscopy provider, Delmic, The Netherlands, and electron microscopy supplier, JEOL to hone and develop several new technologies.
For example, the optical resolution of JEOL's combined light/electron microscope, ClairScope, has been improved while researchers have also developed integrated super-resolution microscopy-SEM for direct correlative light electron microscopy (CLEM).
At the same time, colleagues from Brunel University London have also fabricated novel probes that can be excited with light and electron beams, and used in live cell imaging as well as markers for CLEM.
Meanwhile, in the second project, O'Toole is collaborating with researchers from ptychographic imaging business, Phase Focus, UK, to drive label-free, high contrast, live cell imaging forward.
Ptychography reveals cell states: (a) Live cells imaged at 37°C showing brighter dividing cells (white arrows) and intense lines within the cells marking chromosome alignment prior to cytokinesis (arrowheads). (b) Cells imaged at 40°C showing vesiculation of the cytoplasm due to heat shock (arrows). (c) Apoptotic cell which appears rounder and brighter than the surrounding healthy cells. (d) Cells in a non-confluent preparation showing membrane ruffles (arrows). (e) DIC image of the same field of view as (d) where membrane ruffles are barely visible. From Ptychography – a label free, high-contrast imaging technique for live cells using quantitative phase information. [Marrison et al, Scientific Reports volume 3, Article number: 2369 (2013)]
Phase Focus has developed its 'Virtual Lens' to track live cells, the cell cycle and other vital studies without the need for classic staining or labelling, and recently launched its label-free cell analysis system, which has already been bought by a major pharmaceutical company as well as the Francis Crick Institute and more.
"We've now got this technology proven and I think it's going to be to next big thing that has not yet gone boom," says O'Toole. "The company is right on the edge of where it need to go, and the potential here is monstrous."
Since his arrival at the Bioscience Technology Facility, O'Toole has also collaborated with Instrument heavyweights, Carl Zeiss and Beckman Coulter. Now heading up the entire facility while still leading Imaging and Cytometry, O'Toole is adamant that constructive collaborations are integral to progress and growth.
"For example, you get oversight and first hands-on experiences of new emerging technologies, you get to look at concept products and also beta test," he points out. "But it is also about building networks, communicating to businesses when you have an idea relating to what they are doing, and feeding this back to them in a positive and constructive manner. This in turn can see the resulting product being well suited to the end user applications and needs."
"Collaborations further afield are so very important as you are never going to have all the experts in one location," he adds.
Right now, O'Toole is also Vice President of the Royal Microscopical Society, serves on myriad microscopy-related committees and is heavily involved with microscopy and cytometry training. He largely attributes his success to communication and enthusiasm.
"In any role, you can have good equipment and good staff, and I am incredibly lucky to have excellent staff, but you have to include and inspire your users to use your technology," he says. "At York, the primary aim is to help our researchers achieve high impacting outputs and enable them to access the very latest technologies and applications. By doing this, more and more researchers get involved; it becomes self-perpetuating to a degree. As a core facility and R&D lab, our users often see our lab as an extension of their own, which really makes our team even larger and higher impacting."
"I work with biologists, physicists and chemists, which adds a high degree of diversity that can then be translated to other users," he adds. "So I do think I've just got a great job, and I love it."