Shedding new light on cells with coherent multiphoton nanoscopy
Lead Research Organisation:
Cardiff University
Department Name: School of Biosciences
Abstract
The aim of this research is the realization of a novel imaging modality to enable the observation of living cells and tissues under physiological conditions with unprecedented sensitivity and spatial resolution, without the need to stain them with fluorophores. The technique, based on the interaction of light with matter in the coherent regime, will feature a unique combination two process: Coherent Antistokes Raman Scattering (CARS) of biomolecules in living cells and Four-Wave Mixing (FWM) imaging of metallic nanoparticles (NPs). This technology will progress the field of optical 'nanoscopy', advance our understanding in physics and material sciences, tackle biological problems that are virtually impossible to address with currently available techniques, and will be of relevance in medical applications to improve the diagnostic and treatment of diseases.Optical microscopy is an indispensable tool that is driving progress in cell biology, however most cellular constituents have no colour and they are hard to distinguish under a light microscope unless they are stained. Fluorescence microscopy using organic dyes attached to biomolecules or fluorescent proteins has provided a highly sensitive method of visualizing biomolecules. However, when used for observations in living cells, these modified biomolecules raise questions if their behaviour is real or artefactual. Furthermore, all organic fluorophores are prone to photo-bleaching, an irreversible degradation of the fluorescence intensity after excitation with light, which severely limits time-course observations and is accompanied by toxicity effects and consequent cell damage. In CARS the image contrast is obtained by detecting light which is scattered by vibrating bonds in unstained biomolecules. Although this scattering phenomenon produces a very weak signal, it can be coherently enhanced when two short laser pulses are used to excite the vibrations (generating CARS) so that the scattered light from all bonds of the same type constructively interfere. However, CARS still requires a high number of molecules to achieve sufficient signal for detection, and the existence of a background severely limits its sensitivity. Another problem is the spatial resolution limited by the optical diffraction (>100nm).In this programme, I will overcome these limitations by developing a background free CARS detection combined with the light enhancement occurring in the nanoscale range near a metallic NP, to achieve nanometric spatial resolution and high sensitivity. The NP will be located and tracked using FWM imaging, recently demonstrated in our laboratory, here in a new version to enable nanometric position accuracy in all three directions. FWM detection will also report the local thermal conductivity of the NP surroundings. In addition the microscope will feature trapping of the NP with optical tweezers to position it in a region of interest and/or to measure forces applied to it.The ability to map the intrinsic chemical composition of nanoscale regions together with their thermal and mechanical properties in living cells will have a major impact in solving important biomedical problems. For example, we will determine whether cell membranes perform their function through the assembly of lipid nanodomains or 'rafts'. Their existence is thought to play a key role in basic biological functions and in many diseases (eg influenza and HIV) but is also controversial owing to their small size. Another application will be to determine the local membrane environment associated with endocytosis which is crucial, beyond fundamental biology, for the design of drug delivery and therapeutic strategies. More in general, this novel imaging modality will allow us to address biological systems where the manipulation and photo-toxicity associated with the use of fluorescence markers is unacceptable, e.g. in the areas of in-vitro fertilization and cancer research.
Planned Impact
Who will benefit from this research? The technology developed in this research will progress the field of optical nanoscopy, advance our understanding in physics and material sciences, and tackle biomedical problems that are virtually impossible to address with currently available techniques. We will investigate field enhancement effects in conjunction with coherent nonlinear optics, and, as a first important biological application, we will determine whether cell membranes perform their function through the assembly of lipid nanodomains or 'rafts'. Their existence is thought to play a key role in basic biological functions and in many diseases (eg influenza and HIV) but is also controversial owing to their small size. Another application will be to determine the local membrane environment associated with endocytosis which is crucial, beyond fundamental biology, for the design of drug delivery and therapeutic strategies. Hence this research will impact: - The academic community working in a wide range of disciplines including optical engineering, physics, chemistry, material science, biology and medicine (see Academic beneficiaries section). - The commercial sector, through microscope manufacturers and laser companies interested in the exploitation of the CARS/FWM technology and in the realisation of light sources optimised for it, and drug discovery companies interested in the knowledge of cell membrane dynamics, trafficking and endocytic routes enabled by this technology. - The public sector, through the benefit for public health in the discovery of biological functions which have a key role in diseases. How will they benefit from this research? The contribution of this research to these beneficiaries and to the nation's health, wealth and culture will be mainly through: - Knowledge: via the scientific advancements (on a 3-5 years realistic timescale) in the CARS/FWM technology development and its biological applications. - People: The RA and PhD students on this programme will receive state of the art training in biophotonics. In 2006, Cardiff University established the first MSc Biophotonics course in the UK, jointly taught between the Schools of Physics and Biosciences, sponsored by microscopy-related industrial collaborators. The new lecturer appointed to undertake the teaching duties of the fellow will teach in the course and will collaborate to the programme. This will add to the depth of the research training offered within the course, and will allow better recruiting of excellent students to be trained as Biophotonics leaders of tomorrow. - Improvement of health and quality of life. The direct research outputs will lead to greater understanding of membrane lipid rafts and endocytosis which play a key role in many viral diseases and in drug delivery. Translation of these research outputs into therapeutic strategies will ultimately benefit public health (realistic timescale > 10years). What will be done to ensure that they benefit from this research? I, the RA and the two PhD students in this programme will undertake impact activities as detailed in the Impact plan. Briefly, the team will engage with secondary Schools through the Researchers in Residence Scheme and as STEM ambassadors. Communication with industry and exploitation will occur through an Imaging Symposium annual event, the I-Solve programme, and IP protection routes. To network with the international academic community I will organise an annual series of Biophotonics workshops. I will be specifically trained in communication skills through two Royal Society courses. As part of my professional development as EPSRC leadership fellow I will also be trained in leadership skills through the Leadership and Management Development Programme for Research Team Leaders at Cardiff University. Specialised staff has also been appointed in the form of Innovation and Engagement Officers as detailed in the Impact Plan.
People |
ORCID iD |
| Paola Borri (Principal Investigator) |
Publications
Accanto N
(2012)
Engineering the Spin-Flip Limited Exciton Dephasing in Colloidal CdSe/CdS Quantum Dots
in ACS Nano
Bradley J
(2016)
Quantitative imaging of lipids in live mouse oocytes and early embryos using CARS microscopy.
in Development (Cambridge, England)
Bradley J
(2019)
Dynamic label-free imaging of lipid droplets and their link to fatty acid and pyruvate oxidation in mouse eggs.
in Journal of cell science
Di Napoli C
(2014)
Hyperspectral and differential CARS microscopy for quantitative chemical imaging in human adipocytes.
in Biomedical optics express
Di Napoli C
(2016)
Quantitative Spatiotemporal Chemical Profiling of Individual Lipid Droplets by Hyperspectral CARS Microscopy in Living Human Adipose-Derived Stem Cells.
in Analytical chemistry
Di Napoli C
(2014)
Chemically-specific dual/differential CARS micro-spectroscopy of saturated and unsaturated lipid droplets.
in Journal of biophotonics
Giannakopoulou N
(2020)
Four-wave-mixing microscopy reveals non-colocalisation between gold nanoparticles and fluorophore conjugates inside cells.
in Nanoscale
Karuna A
(2016)
Hyperspectral volumetric coherent anti-Stokes Raman scattering microscopy: quantitative volume determination and NaCl as non-resonant standard.
in Journal of Raman spectroscopy : JRS
Karuna A
(2019)
Label-Free Volumetric Quantitative Imaging of the Human Somatic Cell Division by Hyperspectral Coherent Anti-Stokes Raman Scattering.
in Analytical chemistry
Langbein W
(2018)
Invited Article: Heterodyne dual-polarization epi-detected CARS microscopy for chemical and topographic imaging of interfaces
in APL Photonics
Li B
(2013)
Dual/differential coherent anti-Stokes Raman scattering module for multiphoton microscopes with a femtosecond Ti:sapphire oscillator
in Journal of Biomedical Optics
Masia F
(2013)
Polarization-resolved ultrafast dynamics of the complex polarizability in single gold nanoparticles.
in Physical chemistry chemical physics : PCCP
Masia F
(2013)
Quantitative chemical imaging and unsupervised analysis using hyperspectral coherent anti-Stokes Raman scattering microscopy.
in Analytical chemistry
Masia F
(2018)
Label-free quantitative chemical imaging and classification analysis of adipogenesis using mouse embryonic stem cells
in Journal of Biophotonics
Masia F
(2011)
Exciton dephasing in lead sulfide quantum dots by X -point phonons
in Physical Review B
Masia F
(2015)
Hyperspectral image analysis for CARS, SRS, and Raman data.
in Journal of Raman spectroscopy : JRS
Masia F
(2014)
Sparse sampling for fast hyperspectral coherent anti-Stokes Raman scattering imaging.
in Optics express
Masia F
(2012)
Measurement of the dynamics of plasmons inside individual gold nanoparticles using a femtosecond phase-resolved microscope
in Physical Review B
Masia F
(2012)
Spin-flip limited exciton dephasing in CdSe/ZnS colloidal quantum dots.
in Physical review letters
McPhee CI
(2013)
Measuring the lamellarity of giant lipid vesicles with differential interference contrast microscopy.
in Biophysical journal
Naeem A
(2015)
Giant exciton oscillator strength and radiatively limited dephasing in two-dimensional platelets
in Physical Review B
Nahmad-Rohen A
(2021)
Simultaneous microscopic imaging of thickness and refractive index of thin layers by heterodyne interferometric reflectometry (HiRef)
in Journal of Physics D: Applied Physics
Nahmad-Rohen A
(2020)
Quantitative Label-Free Imaging of Lipid Domains in Single Bilayers by Hyperspectral Coherent Raman Scattering.
in Analytical chemistry
Payne L
(2018)
Wide-Field Imaging of Single-Nanoparticle Extinction with Sub- nm 2 Sensitivity
in Physical Review Applied
Payne L
(2013)
Polarization-resolved extinction and scattering cross-sections of individual gold nanoparticles measured by wide-field microscopy on a large ensemble
in Applied Physics Letters
Payne L
(2015)
Optical micro-spectroscopy of single metallic nanoparticles: quantitative extinction and transient resonant four-wave mixing.
in Faraday discussions
Payne LM
(2021)
Quantitative morphometric analysis of single gold nanoparticles by optical extinction microscopy: Material permittivity and surface damping effects.
in The Journal of chemical physics
Pope I
(2013)
Simultaneous hyperspectral differential-CARS, TPF and SHG microscopy with a single 5 fs Ti:Sa laser.
in Optics express
Pope I
(2014)
Coherent anti-Stokes Raman scattering microscopy of single nanodiamonds.
in Nature nanotechnology
Pope I
(2012)
Live cell imaging with chemical specificity using dual frequency CARS microscopy.
in Methods in enzymology
Pope I
(2023)
Correlative light-electron microscopy using small gold nanoparticles as single probes
in Light: Science & Applications
Pope I
(2023)
Background-free four-wave mixing microscopy of small gold nanoparticles inside a multi-cellular organ
in Applied Physics Letters
Regan D
(2019)
Lipid Bilayer Thickness Measured by Quantitative DIC Reveals Phase Transitions and Effects of Substrate Hydrophilicity.
in Langmuir : the ACS journal of surfaces and colloids
Rocha-Mendoza I
(2012)
Quadruplex CARS micro-spectroscopy
in Journal of Raman Spectroscopy
Slesiona N
(2023)
Correlative Extinction and Single Fluorophore Bleaching Microscopy for Ligand Quantification on Gold Nanoparticles
in Advanced Materials Interfaces
Wagner H
(2013)
Effects of uniaxial pressure on polar whispering gallery modes in microspheres
in Journal of Applied Physics
Wang Y
(2021)
A primary effect of palmitic acid on mouse oocytes is the disruption of the structure of the endoplasmic reticulum.
in Reproduction (Cambridge, England)
Wang Y
(2022)
Quantitatively linking morphology and optical response of individual silver nanohedra.
in Nanoscale
| Title | Contemporary Dance OPTO-NANO |
| Description | The professional dancer and choreographer Jack Philp created a dance piece inspired by the research in my laboratory, with funding from the Arts Council Wales. This resulted in a showcase performance at the National dance House in Cardiff in November 2019 and in a video posted in social media. |
| Type Of Art | Performance (Music, Dance, Drama, etc) |
| Year Produced | 2019 |
| Impact | The work has attracted considerable interest in social media. Several members of the general public attended the performance showcase which was followed by a question and answer session in which I had the opportunity to explain my research to the wider public. With the coreographer we plan to bring this forward into a dance tour and we are seeking further funding. |
| URL | http://www.jackphilpdance.co.uk/opto-nano |
| Description | The aim of this research was the realization of a novel optical imaging modality to enable the observation of living cells and tissues under physiological conditions with unprecedented sensitivity and spatial resolution, without the need to stain them with fluorophores. The idea of the technique was to use two coherent light-matter interaction effects, and their combination, namely Coherent Antistokes Raman Scattering (CARS) microscopy of endogenous biomolecules in living cells and resonant Four-Wave Mixing (FWM) imaging of metallic nanoparticles (NPs), each featuring many advantages compared to existing methods. Within this research programme we have: 1) Developed a novel heterodyne-CARS (H-CARS) microscope for background-free detection. 2) Demonstrated FWM detection of NPs with nanometric position accuracy in 3D 3) Integrated H-CARS and FWM detection modalities in the same instrument 4) Demonstrated detection of CARS locally-enhanced in the vicinity of a metallic NP 5) Demonstrated FWM detection with nanoparticles of different shapes and sizes 6) Demonstrated FWM detection of single NPs background free inside cells in 3D, correlatively with confocal fluorescence of labelled ligands. |
| Exploitation Route | These findings have already resulted in a large amount of international research papers (> 40) and conference presentations since 2011, with more articles being in preparation and expected to be published over the coming year. The research trend of surface-enhanced CARS is very promising and the intention is to pursue it with future grant applications, to explore basic questions in biophysics. From a non-academic impact point of view, both CARS and FWM of NPs are promising imaging modality for applications in drug delivery, and we have specific contacts with pharmaceutical and biotech companies (GSK, Cellesce) to explore the commercial impact of the technology. We also have ongoing collaborations with major NP manufacturers (BBI solutions, NanoComposix, Sona Nanotech). We are now actively working to establish a knowledge transfer partnership with manufacturers of analytical instrumentation. We note that the FWM imaging invention has been protected with a filed patent application. |
| Sectors | Chemicals,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology |
| Description | Key findings of this research have been used to strengthen collaborations with particle manufacturer companies (BBI Solutions, Nanocomposix, Sona Nanotech) and pharmaceutical and biotech companies (GSK, Cellesce). We have had two iCASE studentships with these companies. Plans are on going to establish a knowledge transfer project with companies manufacturing analytical instrumentation. We are also working toward demonstrating FWM imaging as a novel tool for correlative light electron microscopy in collaboration with Bristol University and microscope companies. We note that our FWM imaging invention has been protected with a filed patent application. In 2019 we filed a second patent application to protect the invention of measuring accurately the size and shape individual nanoparticles by wide-field transmission microscopy. Within this project we also actively engaged in outreach activities to the wider public, and showcased our work at the Cardiff Science Festival in 2014. In 2019 an outreach project with the arts was commenced, resulting in a contemporary dance choreography being created, inspired by the work from this research, and showcased at the National Dance Company House in Cardiff. |
| First Year Of Impact | 2011 |
| Sector | Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology |
| Impact Types | Cultural,Economic |
| Description | BEIS/ Universities UK Researcher roundtable in Cardiff about the impact of Brexit on international research collaborations and bidding for funding for internationally collaborative research projects |
| Geographic Reach | Europe |
| Policy Influence Type | Contribution to a national consultation/review |
| Description | A novel imaging approach to track brain cancer metabolism - ISSF3 Collaboration Panel: Cross-Disciplinary Award, Cardiff University |
| Amount | £22,545 (GBP) |
| Organisation | Wellcome Trust |
| Sector | Charity/Non Profit |
| Country | United Kingdom |
| Start | 09/2019 |
| End | 04/2020 |
| Description | Creating super-scattering Raman-active genetically encoded proteins |
| Amount | £198,490 (GBP) |
| Funding ID | EP/V048147/1 |
| Organisation | Engineering and Physical Sciences Research Council (EPSRC) |
| Sector | Public |
| Country | United Kingdom |
| Start | 01/2021 |
| End | 06/2022 |
| Description | Experimental Equipment Call |
| Amount | £731,951 (GBP) |
| Funding ID | EP/M028313/1 |
| Organisation | Engineering and Physical Sciences Research Council (EPSRC) |
| Sector | Public |
| Country | United Kingdom |
| Start | 04/2015 |
| End | 03/2016 |
| Description | ISSF3 Cross-Disciplinary Award: Uncovering metabolic errors in foetal brain development through advanced lipid imaging |
| Amount | £37,067 (GBP) |
| Organisation | Wellcome Trust |
| Sector | Charity/Non Profit |
| Country | United Kingdom |
| Start | 11/2020 |
| End | 05/2021 |
| Description | Initial Training Networks Call identifier: FP7-PEOPLE-2013-ITN |
| Amount | € 584,513 (EUR) |
| Funding ID | 607842 |
| Organisation | European Commission |
| Sector | Public |
| Country | European Union (EU) |
| Start | 10/2013 |
| End | 09/2017 |
| Description | Life Sciences Bridging Fund |
| Amount | £5,000 (GBP) |
| Funding ID | LSBF/R4-004 |
| Organisation | Government of Wales |
| Sector | Public |
| Country | United Kingdom |
| Start | 08/2016 |
| End | 10/2016 |
| Description | MR/X018318/1 - Proteostatic regulation of glioblastoma stemness - MRC |
| Amount | £1,299,880 (GBP) |
| Funding ID | MR/X018318/1 |
| Organisation | Medical Research Council (MRC) |
| Sector | Public |
| Country | United Kingdom |
| Start | 06/2023 |
| End | 05/2027 |
| Description | Measuring the sp3/sp2 Carbon Content Ratio in a Single Nanodiamond Using Quantitative Optical Microscopy |
| Amount | £80,000 (GBP) |
| Funding ID | EPSRC CDT Diamond Science and Technology |
| Organisation | Cardiff University |
| Sector | Academic/University |
| Country | United Kingdom |
| Start | 10/2019 |
| End | 09/2022 |
| Description | Multiphoton Microscopy and Ultrafast Spectroscopy: Imaging meets Quantum (MUSIQ) |
| Amount | € 4,034,447 (EUR) |
| Funding ID | 812922 |
| Organisation | Marie Sklodowska-Curie Actions |
| Sector | Charity/Non Profit |
| Country | Global |
| Start | 04/2019 |
| End | 03/2023 |
| Description | Non-invasive chemically-specific imaging of lung inflammation |
| Amount | £24,984 (GBP) |
| Funding ID | UKRI Technology Touching Life IBIN |
| Organisation | King's College London |
| Sector | Academic/University |
| Country | United Kingdom |
| Start | 05/2021 |
| End | 10/2021 |
| Description | Royal Society Challenge Grant Awards |
| Amount | £91,269 (GBP) |
| Funding ID | CH160031 |
| Organisation | The Royal Society |
| Sector | Charity/Non Profit |
| Country | United Kingdom |
| Start | 12/2016 |
| End | 12/2017 |
| Description | Science Committee - Multidisciplinary Project Award |
| Amount | £403,597 (GBP) |
| Funding ID | C368/A22099 |
| Organisation | Cancer Research UK |
| Sector | Charity/Non Profit |
| Country | United Kingdom |
| Start | 08/2016 |
| End | 07/2019 |
| Description | TTL Integrated Biological Imaging Network Pump-priming Project: A novel correlative light electron microscopy (CLEM) technique to unravel the nano-toxicology of single gold nanoparticles in terrestrial isopods |
| Amount | £24,808 (GBP) |
| Organisation | United Kingdom Research and Innovation |
| Department | Technology Touching Life |
| Sector | Public |
| Country | United Kingdom |
| Start | 10/2019 |
| End | 03/2020 |
| Description | Tracking the motion of single nanoparticles inside living cells: New insights into intracellular crowdedness |
| Amount | £80,000 (GBP) |
| Funding ID | EPSRC DTP account |
| Organisation | Cardiff University |
| Sector | Academic/University |
| Country | United Kingdom |
| Start | 01/2021 |
| End | 06/2024 |
| Description | Training Grant, Industrial CASE with BBI |
| Amount | £94,126 (GBP) |
| Funding ID | BB/L015889/1 |
| Organisation | Biotechnology and Biological Sciences Research Council (BBSRC) |
| Sector | Public |
| Country | United Kingdom |
| Start | 10/2014 |
| End | 09/2018 |
| Description | Unravelling the dynamics of lipid bilayers and membrane proteins and their reaction to toxins and viral infection on the single molecule level using label-free microscopy |
| Amount | £80,000 (GBP) |
| Funding ID | EPSRC DTP account |
| Organisation | Cardiff University |
| Sector | Academic/University |
| Country | United Kingdom |
| Start | 10/2020 |
| End | 03/2024 |
| Description | Welsh Government Award - LIFE SCIENCES BRIDGING FUND Optical Nanosizer: Wide-field extinction micro-spectroscopy for nanoparticle analysis |
| Amount | £47,894 (GBP) |
| Funding ID | LSBF/R6-005 |
| Organisation | Government of Wales |
| Sector | Public |
| Country | United Kingdom |
| Start | 06/2017 |
| End | 03/2018 |
| Description | BBI-nanoparticle analysis |
| Organisation | BBI Solutions |
| Country | United Kingdom |
| Sector | Private |
| PI Contribution | We have developed a novel method to quantify the absorption and scattering cross section of single metallic nanoparticles using wide-field microscopy. The method enables simultaneous acquisition of hundreds of nanoparticles for statistical analysis. The method is simple, rapid, and quantitative. Precise characterisation of nanoparticle sizes and shapes is key to manufacturing quality control. Current techniques for particle characterisation are dynamics light scattering (DLS), which is easy to use but not very accurate and provides only the effective mean size (hydrodynamic radius), and transmission electron microscopy (TEM) which is time consuming and expensive. The optical extinction microscopy method developed by us overcomes these limitations and effectively complements these analysis techniques. BBI Solutions which is a world leading manufacturer of gold nanoparticles for lateral flow analysis and in-vitro diagnostics (http://www.bbisolutions.com/). They have a major nanoparticle synthesis and analysis headquarter at Cardiff. They see huge potential benefit in the use of optical extinction for particle analysis and quality control, owing to the accuracy and versatility of the method which can be implemented cost-effectively on a conventional wide-field microscope and is applicable to any nanoparticle. |
| Collaborator Contribution | BBI Solutions has provided us with nanoparticle materials, including custom made, and access to their analysis tools. They have dedicated a significant amount of time in technical discussions with us. They have been industrial partners on an EPSRC funded project, and are co-investigators in a BBSRC iCASE studentship with us where they have hosted the PhD student on-site for a significant amount of time. |
| Impact | This collaboration was aimed at providing BBI solutions with a cost-effective tool for particle analysis which they can use instead of costly TEM, hence having a clear financial benefit. We also explored with them ways in which they could expand their portfolio of nanoparticle manufacturing, using our quantitative analysis to provide better product specification to customers and improve nanoparticle design. |
| Start Year | 2013 |
| Description | CRS on organoids with Cellesce |
| Organisation | Cellesce Ltd |
| Country | United Kingdom |
| Sector | Private |
| PI Contribution | Expertise in optical microscopy technology developments beyond state of the art, in particular label-free microscopy based on Coherent Raman Scattering, and associated quantitative image analysis. Application of CRS microscopy to organoid samples provided by the company. |
| Collaborator Contribution | Cellesce has a patented technology for the expansion of human-derived "organoids" at scale for high-throughput screening in the pharmaceutical industry. Organoids are 3D clusters of cells, derived from patient biopsies, typically from tumours, that accurately represent the architecture and physiology of the original tissue. When grown on a commercial scale, organoids represent a new, improved and transformational, model for drug discovery and for clinical and academic research. In this partnership, Cellesce is providing us with living and fixed organoids samples, to be investigated with our optical microscopy technologies beyond state of the art. |
| Impact | Presentations at conferences and meetings. New grant applications. The collaboration is multi-disciplinary at the physics-life sciences interface. |
| Start Year | 2019 |
| Description | GSK-CARS |
| Organisation | GlaxoSmithKline (GSK) |
| Country | Global |
| Sector | Private |
| PI Contribution | We have developed a chemically specific label-free imaging method of living cells, with associated quantitative image analysis. GSK is interested in exploring how the method can be used for high-content image analysis in the context of drug discovery and delivery. |
| Collaborator Contribution | GSK is our industrial partner in a iCASE PhD studentship. They have committed their time and access to their resources, in terms of drug formulations, cell lines and analytical methods. |
| Impact | This collaboration is multi-disciplinary involving physics, biochemistry and molecular cell biology. |
| Start Year | 2014 |
| Title | ANALYSING NANO-OBJECTS |
| Description | Methods and apparatus for analysis of nano-objects using wide-field bright field transmission techniques are described. Such methods may comprise acquiring a plurality of images of a sample (124) comprising a plurality of nano-objects using bright field illumination via a continuously variable spectral filter (114), and identifying a nano-object within the sample in the plurality of images, wherein the position of the nano-object changes between images. Using data extracted from the plurality of images, an extinction cross-section of the identified nano-object may be quantitatively determined. |
| IP Reference | WO2019020975 |
| Protection | Patent application published |
| Year Protection Granted | 2019 |
| Licensed | No |
| Impact | This development has resulted in a number of research papers that have been published in high quality international journals. We are seeking to engage with companies manufacturing analytical instrumentation to licence the IP. |
| Title | SURFACE PLASMON FOUR-WAVE MIXING MICROSCOPY |
| Description | Laser pulses are applied to surface plasmon resonant articles such as gold nanoparticles within a microscopy sample to generate a four-wave mixing signal that is detected as the output of the microscopy process. |
| IP Reference | WO2010070337 |
| Protection | Patent granted |
| Year Protection Granted | 2010 |
| Licensed | No |
| Impact | Collaboration with nanoparticle manufacturers and microscope companies |
| Title | Heterodyne CARS microscopy |
| Description | We have developed a novel label-free technique based on chemical imaging via coherent anti-Stokes Raman scattering microscopy using a dual-polarization balanced heterodyne detection in epi-geometry (eH-CARS). This technique provides a unique combination of background-free chemically-specific image contrast for nanoparticles and interfaces, shot-noise limited detection, and phase sensitivity. |
| Type Of Technology | New/Improved Technique/Technology |
| Year Produced | 2015 |
| Impact | This new technique could prove instrumental to investigate the chemical composition of thin films with high spatial resolution, topographic sensitivity, and label-free. It could be a powerful tool to study single lipid bilayers, including model and cellular membranes, as well as single nanoparticles. |
| Title | Hyper spectral image analysis software |
| Description | The software developed is an image analysis tool to quantitatively retrieve concentrations of chemical components and spectral Raman-like profiles from hyperspectral CARS microscopy data |
| Type Of Technology | Software |
| Year Produced | 2013 |
| Impact | Quantitative determination of concentration of chemical components (lipids, proteins, DNA) and their spatial distribution in cells and tissues from label-free hyperspectral CARS microscopy data. The software has been made freely available, and has been utilized by several groups. It is also applicable beyond CARS data, to any dataset that requires a decomposition analysis (eg fluorescence spectra). It is becoming part of an image analysis suite embedded in an institutional image facility. |
| URL | http://langsrv.astro.cf.ac.uk/HIA/HIA.html |
| Title | Quantitative Extinction analysis |
| Description | We have developed an optical extinction microscopy analysis method to quantify the optical extinction cross-section s_ext of single nanoparticles using wide-field microscopy. The method utilities the simultaneous acquisition of hundreds of nanoparticles for statistical analysis. The method is simple, rapid, and quantitative. We have demonstrated a sensitivity of s_ext below 1nm^2 corresponding to the detection of a single gold nanoparticle of 2nm diameter, using only ~1min total acquisition time. From the measurement of s_ext and its dependence on the light polarization, an accurate quantification of particle sizes and asymmetries at the single particle level is rapidly obtained for a statistically-relevant number (>100) of particles. An Extinction Suite Macro for ImageJ was also developed, for quantitative data analysis. |
| Type Of Technology | New/Improved Technique/Technology |
| Year Produced | 2013 |
| Impact | This technique has generated commercial interested with our industrial collaborators BBI Solution, a world leading nanoparticle manufacturer. Precise characterisation of nanoparticle sizes and shapes is key to manufacturing quality control. Current techniques for particle characterisation are dynamics light scattering (DLS), which is easy to use but not very accurate and provides only the effective mean size (hydrodynamic radius), and transmission electron microscopy (TEM) which is time consuming and expensive. The optical extinction microscopy method developed by us overcomes these limitations and effectively complements these analysis techniques. Malvern Panalytical Ltd manufacturing analytical instrumentation including DLS, is interested in a knowledge transfer partnership on this technology. |
| URL | http://langsrv.astro.cf.ac.uk/Crosssection/Extinction_Suite/Extinction_Suite.html |
| Title | Resonant FWM imaging |
| Description | We have developed a novel optical microscopy technique to visualise small single metallic nanoparticels (NPs), without the need of any fluorophore labelling, in a very specific and background free way, even in highly scattering environments such as cells and tissues. In this method individual small (radii below 20nm) metallic NPs are visualised with high contrast and photo-stability, free from background and at powers corresponding to negligible photothermal heating, via their coherent nonlinear optical emission called Four-Wave Mixing (FWM) in resonance with the NP surface plasmon. The technique has been implemented to work both in transmission and reflection geometry. It includes a polarisation resolved analysis to detect particle asymmetry, a phase resolved analysis for biosensing, and a time-resolved analysis to quantify thermal and mechanical couplings of NPs with their environments. |
| Type Of Technology | New/Improved Technique/Technology |
| Year Produced | 2012 |
| Impact | The technique is being utilized extensively over several research projects, including a collaborative project with Bristol University with interest in correlative light-electron microscopy . It is planned to become part of an imaging facility at Cardiff University. |
| URL | http://www.cardiff.ac.uk/people/view/81122-borri-paola |
| Title | Single particle tracking via four-wave-mixing optical vortex interferometry |
| Description | We have developed and demonstrated a new four-wave-mixing interferometry technique, whereby the position of a single nonfluorescing gold nanoparticle is determined with nanometric precision in 3D from rapid single-point measurements at 1-ms acquisition time by exploiting optical vortices. The technique is also uniquely sensitive to particle asymmetries of only 0.5% ellipticity, corresponding to a single atomic layer of gold, as well as particle orientation. This method opens new ways of unraveling single-particle trafficking within complex 3D architectures. |
| Type Of Technology | New/Improved Technique/Technology |
| Year Produced | 2015 |
| Impact | Ultimately, this new method paves the way towards a new form of single-particle tracking, where not only the nanoparticle position but also its asymmetry, orientation, and chirality are detected with submillisecond time resolution, revealing much more information about the nanoparticle and its complex dynamics (e.g., hindered rotation) while moving and interacting within a disordered 3D environment. As such it can have many applications in physical science systems (e.g. diffusion in complex liquids and gels, liquid crystals) and in the life sciences (single-molecule intra-cellular trafficking). |
| Title | quantitative DIC microscopy |
| Description | We have developed an analysis technique to quantify the thickness of thin transparent objects using optical microscopy, based on quantitative differential interference contrast. The method was demonstrated on giant unilamellar lipid vesicles. It is generally applicable to many other structures, for example nanoparticles, thin films and waveguides. |
| Type Of Technology | New/Improved Technique/Technology |
| Year Produced | 2013 |
| Impact | Since its introduction the technique has been widely used (by our and other labs) to 1) measure the volume of transparent particles, such as nanodiamonds 2) measure the thickness of lipid bilayers both suspended and surface attached 3) measure the thickness of thin dielectric layers for waveguide design and processing |
| URL | http://dx.doi.org/10.1016/j.bpj.2013.07.048 |
| Description | BioNanoPhotonics Symposium |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | International |
| Primary Audience | Industry/Business |
| Results and Impact | This is a scientific symposium that I organise every two years, with international invited speakers and participation from industry and postgraduate students. We started having 40 delegates in 2011 and reached 80 delegates in 2015. The event has sparked many scientific discussions and further collaboration with industry among the delegates. |
| Year(s) Of Engagement Activity | 2011,2013,2015,2017 |
| URL | http://www.bionanophoto.org.uk/ |
| Description | Cardiff Science Festival 2014 |
| Form Of Engagement Activity | A talk or presentation |
| Part Of Official Scheme? | No |
| Geographic Reach | Regional |
| Primary Audience | Public/other audiences |
| Results and Impact | A stand showcasing our group research activity to the general public was set-up at the Cardiff Science Festival on 19 -20 July 2014. At this stand four members of our group (two postdoctoral researchers and two PhD students) gave practical demonstrations and gifts, and presented an "attention grabbing" poster based around the concept of "Blinged Nanoparticles" |
| Year(s) Of Engagement Activity | 2014 |
| URL | https://www.facebook.com/Cardiff-Science-Festival-386755174684198/ |
| Description | Contemporary Dance project OPTO NANO |
| Form Of Engagement Activity | Participation in an activity, workshop or similar |
| Part Of Official Scheme? | No |
| Geographic Reach | Regional |
| Primary Audience | Public/other audiences |
| Results and Impact | In collaboration with a contemporary dance professional choreographer Jack Philp, an outreach project was started aimed at creating a dance piece inspired by the biophotonics research in my lab. Jack was awarded a grant from the Arts Council Wales to work on the creation of the piece which was showcased at an open rehearsal to the public at the National Dance House in Cardiff in November 2019. The work is titled OPTO-NANO and resulted in a promotional video and posting on social media. Jack is working toward expanding the work into a tour and we are seeking further funding. |
| Year(s) Of Engagement Activity | 2019 |
| URL | http://www.jackphilpdance.co.uk/opto-nano |