Dr Iestyn Pope
Professional Specialist
Overview
Research overview
I am a member of the BioPhotonics & Quantum Optoelctronics Group of Prof Paola Borri (School of Biosciences) and Prof Wolfgang Langbein (School of Physics & Astronomy). As part of this group who's interests lie at the interface between life and physical sciences I am involved in the following research activities:
Development of a new generation single source laser-scanning multiphoton microscope, capable of simultaneously acquiring differential-Coherent anti-Stokes Raman Scattering (D-CARS), Two Photon Fluorescence (TPF) and Second Harmonic Generation (SHG) images for studies on living cells. Using this new microscope, we are probing the following biological areas:
- The uptake of different types of lipid compounds (e.g. saturated / unsaturated) to determine the dynamics of lipid droplet formation and development in living cells.
- The study of the change in distribution of lipid droplets during oocyte development.
- The use of hyperspectral CARS data sets to generate chemical maps of cells and organoid structures.
- The use of nanodiamonds as optical labels and contrast agents in live cells.
The use of resonant Four-Wave Mixing (FWM) to image gold and silver nanoparticles inside cells.
Links
Publication
2023
- Slesiona, N., Payne, L., Pope, I., Borri, P., Langbein, W. and Watson, P. 2023. Correlative extinction and single fluorophore bleaching microscopy for ligand quantification on gold nanoparticles. Advanced Materials Interfaces 10(24), article number: 2300568. (10.1002/admi.202300568)
- Pope, I., Ferreira, N. G. C., Kille, P., Langbein, W. and Borri, P. 2023. Background-free four-wave mixing microscopy of small gold nanoparticles inside a multi-cellular organ. Applied Physics Letters 122, article number: 153701. (10.1063/5.0140651)
- Pope, I. et al. 2023. Correlative light-electron microscopy using small gold nanoparticles as single probes. Light: Science & Applications 12, article number: 80. (10.1038/s41377-023-01115-4)
2022
- Wang, Y., Pope, I., Brennan-Craddock, H., Poole, E., Langbein, W., Borri, P. and Swann, K. 2022. A primary effect of palmitic acid on mouse oocytes is the disruption of the structure of the endoplasmic reticulum. Reproduction 163(1), pp. 45-56. (10.1530/REP-21-0332)
2021
- Boorman, D., Pope, I., Masia, F., Langbein, W., Hood, S., Borri, P. and Watson, P. 2021. Hyperspectral CARS microscopy and quantitative unsupervised analysis of deuterated and non-deuterated fatty acid storage in human cells. The Journal of Chemical Physics 155(22), article number: 224202. (10.1063/5.0065950)
- Boorman, D., Pope, I., Masia, F., Watson, P., Borri, P. and Langbein, W. 2021. Quantification of the nonlinear susceptibility of the hydrogen and deuterium stretch vibration for biomolecules in coherent Raman microspectroscopy. Journal of Raman Spectroscopy 52(9), pp. 1540-1551. (10.1002/jrs.6164)
- Pope, I. et al. 2021. Background-free 3D four-wave mixing microscopy of single gold nanoparticles inside biological systems. Presented at: European Conference on Biomedical Optics 2021, Munich, Germany, 20-24 June 2021European Conferences on Biomedical Optics 2021 (ECBO). OSA Technical Digest Optical Society of America pp. EM3B.6.
- Pope, I. et al. 2021. Identifying subpopulations in multicellular systems by quantitative chemical imaging using label-free hyperspectral CARS microscopy. Analyst 146(7), pp. 2277-2291. (10.1039/D0AN02381G)
2020
- Nahmad-Rohen, A., Regan, D., Masia, F., McPhee, C., Pope, I., Langbein, W. and Borri, P. 2020. Quantitative label-free imaging of lipid domains in single bilayers by hyperspectral coherent Raman scattering. Analytical Chemistry 92(21) (10.1021/acs.analchem.0c03179)
- Giannakopoulou, N. et al. 2020. Four-wave-mixing microscopy reveals non-colocalisation between gold nanoparticles and fluorophore conjugates inside cells. Nanoscale 12(7), pp. 4622-4635. (10.1039/C9NR08512B)
- Lloyd, D. et al. 2020. Functional imaging of a model unicell: Spironucleus vortens as an anaerobic but aerotolerant flagellated protist. Advances in Microbial Physiology 76, pp. 41-79. (10.1016/bs.ampbs.2020.01.002)
2019
- Bradley, J., Pope, I., Wang, Y., Langbein, W., Borri, P. and Swann, K. 2019. Dynamic label-free imaging of lipid droplets and their link to fatty acid and pyruvate oxidation in mouse eggs. Journal of Cell Science 132(13), article number: jcs228999. (10.1242/jcs.228999)
- Borri, P., Giannakopoulou, N., Zoriniants, G., Pope, I., Masia, F., Watson, P. and Langbein, W. 2019. Imaging and tracking single plasmonic nanoparticles in 3D background-free with four-wave mixing interferometry. Presented at: SPIE BIOS, San Francisco, CA, USA, 2-7 February 2019Proceedings Volume 10894, Plasmonics in Biology and Medicine XVI, Vol. 108940. Society of Photo-Optical Instrumentation Engineers (SPIE) pp. 34., (10.1117/12.2507618)
- Boorman, D., Pope, I., Langbein, W., Hood, S., Borri, P. and Watson, P. 2019. Optimisation of multimodal coherent anti-Stokes Raman scattering microscopy for the detection of isotope-labelled molecules. Presented at: SPIE BIOS, San Francisco, CA, USA, 2-7 February 2019Proceedings Volume 10890, Label-free Biomedical Imaging and Sensing (LBIS) 2019, Vol. 108900. Society of Photo-Optical Instrumentation Engineers (SPIE) pp. 4., (10.1117/12.2509280)
- Langbein, W., Harlow, D. S., Regan, D., Pope, I. and Borri, P. 2019. Heterodyne dual-polarization epi-detected CARS microscopy for chemical and topographic imaging of interfaces. Presented at: SPIE BIOS, San Francisco, California, US, 2-7 Feb 2019Label-free Biomedical Imaging and Sensing (LBIS) 2019, Vol. 10890. Bellingham, Washington: Society of Photo-optical Instrumentation Engineers (SPIE), (10.1117/12.2507636)
- Borri, P., Bradley, J., Pope, I., Langbein, W. and Swann, K. 2019. Imaging lipids in living mammalian oocytes and early embryos by coherent Raman scattering microscopy. Presented at: SPIE BIOS, San Francisco, CA, USA, 2-7 February 2019Proceedings Volume 10890, Label-free Biomedical Imaging and Sensing (LBIS) 2019, Vol. 108900. Society of Photo-Optical Instrumentation Engineers (SPIE) pp. 3., (10.1117/12.2506248)
2018
- Pope, I. et al. 2018. Coherent Raman Scattering microscopy: technology developments and biological applications. Presented at: 20th International Conference on Transparent Optical Networks (ICTON), Bucharest, Romania, 1-5 Jul 201820th International Conference on Transparent Optical Networks (ICTON). IEEE, (10.1109/ICTON.2018.8473706)
- Borri, P., Zorinyants, G., Giannakopoulou, P., Masia, F., Pope, I. and Langbein, W. 2018. Imaging and tracking single plasmonic nanoparticles in 3D background-free with four-wave mixing interferometry. Presented at: International Conference on Transparent Optical Networks, Bucharest, Romania, 1-5 Jul 201820th International Conference on Transparent Optical Networks (ICTON). IEEE, (10.1109/ICTON.2018.8473874)
- Langbein, W., Regan, D., Pope, I. and Borri, P. 2018. Heterodyne dual-polarization epi-detected CARS microscopy for chemical and topographic imaging of interfaces. APL Photonics 3, article number: 92402. (10.1063/1.5027256)
- Masia, F., Pope, I., Watson, P., Langbein, W. and Borri, P. 2018. Bessel-beam hyperspectral CARS microscopy with sparse sampling: enabling high-content high-throughput label-free quantitative chemical imaging. Analytical Chemistry 90(6), pp. 3775-3785. (10.1021/acs.analchem.7b04039)
2016
- Bradley, J., Pope, I., Masia, F., Sanusi, R., Langbein, W., Swann, K. and Borri, P. 2016. Quantitative imaging of lipids in live mouse oocytes and early embryos using CARS microscopy. Development 143, pp. 2238-2247. (10.1242/dev.129908)
- Di Napoli, C., Pope, I., Masia, F., Langbein, W., Watson, P. and Borri, P. 2016. Quantitative spatiotemporal chemical profiling of individual lipid droplets by hyperspectral CARS microscopy in living human adipose-derived stem cells. Analytical Chemistry 88(7), pp. 3677-3685. (10.1021/acs.analchem.5b04468)
2014
- Pope, I. et al. 2014. Coherent anti-Stokes Raman scattering microscopy of single nanodiamonds. Nature Nanotechnology 9(11), pp. 940-946. (10.1038/nnano.2014.210)
- Di Napoli, C., Pope, I., Masia, F., Watson, P. D., Langbein, W. W. and Borri, P. 2014. Hyperspectral and differential CARS microscopy for quantitative chemical imaging in human adipocytes. Biomedical Optics Express 5(5), pp. 1378-1390. (10.1364/BOE.5.001378)
- Di Napoli, C., Masia, F., Pope, I., Otto, C., Langbein, W. W. and Borri, P. 2014. Chemically-specific dual/differential CARS micro-spectroscopy of saturated and unsaturated lipid droplets. Journal of Biophotonics 7(1-2), pp. 68-76. (10.1002/jbio.201200197)
2013
- Pope, I., Langbein, W. W., Watson, P. D. and Borri, P. 2013. Simultaneous hyperspectral differential-CARS, TPF and SHG microscopy with a single 5 fs Ti:Sa laser. Optics Express 21(6), pp. 7096-7106. (10.1364/OE.21.007096)
2012
- Pope, I., Langbein, W. W., Borri, P. and Watson, P. D. 2012. Live cell imaging with chemical specificity using dual frequency CARS microscopy. In: Conn, P. M. ed. Imaging and Spectroscopic Analysis of Living Cells — Optical and Spectroscopic Techniques. Methods in Enzymology Vol. 504. Amsterdam: Elsevier, pp. 273-291., (10.1016/B978-0-12-391857-4.00014-8)
2011
- Pope, I., Barber, P. R., Horn, S., Ainsbury, E., Rothkamm, K. and Vojnovic, B. 2011. A portable microfluidic fluorescence spectrometer device for γ-H2AX-based biological dosimetry. Radiation Measurements 46(9), pp. 907-911. (10.1016/j.radmeas.2011.02.004)
- Langbein, W. W., Rocha-Mendoza, I., Masia, F., Di Napoli, C., Pope, I., Watson, P. D. and Borri, P. 2011. Differential CARS microscopy with linearly chirped femtosecond laser pulses. Presented at: Multiphoton Microscopy in the Biomedical Sciences XI, San Francisco, CA, USA, 23-25 January 2011 Presented at Periasamy, A., König, K. and So, P. T. C. eds.Multiphoton Microscopy in the Biomedical Sciences XI, Vol. 7903. SPIE Proceedings SPIE pp. 79031I., (10.1117/12.873872)
- Pope, I., Langbein, W. W., Borri, P. and Watson, P. D. 2011. CARS imaging for high throughput microscopy. Biotech International 23(Apr/Ma), pp. 11-13.
- Flaccavento, G., Lempitsky, V., Pope, I., Barber, P., Zisserman, A., Noble, J. A. and Vojnovic, B. 2011. Learning to count cells: Applications to lens-free imaging of large fields. Presented at: Microscopic Image Analysis with Applications in Biology, Heidelberg, Germany, 2 Sept 2011.
2008
- Vojnovic, B., Barber, P., Pope, I., Smith, P. J. and Errington, R. J. 2008. Detecting objects. WO/2008/090330 [Patent].
2007
- Njoh, K. et al. 2007. Live cell tracking on an optical biochip platform. Presented at: Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V, San Jose, CA, USA, 20 January 2007 Presented at Farkas, D. L., Leif, R. C. and Nicolau, D. V. eds.Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V. Proceedings of SPIE Vol. 6441. Bellingham, WA: SPIE pp. 64410X., (10.1117/12.698935)
- Morris, D. et al. 2007. Development of an optical biochip for the analysis of cell environment sensitivity. Presented at: Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V, San Jose, CA, USA, 20 January 2007 Presented at Farkas, D. L., Leif, R. C. and Nicolau, D. V. eds.Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V. Proceedings of SPIE Vol. 6441. Bellingham, WA: SPIE pp. 64410V., (10.1117/12.700052)
- Matthews, D. R. et al. 2007. A fluorescence biochip with a plasmon active surface. Presented at: Plasmonics in Biology and Medicine IV, San Jose, CA, USA, 20 January 2007 Presented at Vo-Dinh, T. and Lakowicz, J. R. eds.Plasmonics in Biology and Medicine IV. Proceedings of SPIE Vol. 6450. Bellingham, WA: SPIE pp. 645006., (10.1117/12.698948)
2006
- Thomson, J. D. et al. 2006. The influence of acceptor anneal temperature on the performance of InGaN/GaN quantum well light-emitting diodes. Journal of Applied Physics 99(2), article number: 24507. (10.1063/1.2165405)
2005
- Brown, I. H. et al. 2005. Determination of the Piezoelectric Field in InGaN Quantum Wells. Applied Physics Letters 86, article number: 131108. (10.1063/1.1896446)
- Thomson, J. D. et al. 2005. The role of acceptor anneal temperature on the performance of InGaN/GaN quantum well light emitting diodes. Presented at: Physics and simulation of optoelectronic devices XIII, San Jose, USA, 24-27 January 2005 Presented at Osinski, M., Henneberger, F. and Amano, H. eds.Physics and Simulation of Optoelectronic Devices XIII. Proceedings of SPIE Vol. 5722. Bellingham: SPIE pp. 425-430., (10.1117/12.591897)
2004
- Pope, I. 2004. The characterisation of InGaN/GaN quantum well light emitting diodes. PhD Thesis, Cardiff University.
2003
- Pope, I., Smowton, P. M., Blood, P., Thomson, J. D., Kappers, M. J. and Humphreys, C. J. 2003. Carrier leakage in InGaN quantum well light-emitting diodes emitting at 480 nm. Applied Physics Letters 82(17), pp. 2755-2757. (10.1063/1.1570515)
Adrannau llyfrau
- Pope, I., Langbein, W. W., Borri, P. and Watson, P. D. 2012. Live cell imaging with chemical specificity using dual frequency CARS microscopy. In: Conn, P. M. ed. Imaging and Spectroscopic Analysis of Living Cells — Optical and Spectroscopic Techniques. Methods in Enzymology Vol. 504. Amsterdam: Elsevier, pp. 273-291., (10.1016/B978-0-12-391857-4.00014-8)
Cynadleddau
- Pope, I. et al. 2021. Background-free 3D four-wave mixing microscopy of single gold nanoparticles inside biological systems. Presented at: European Conference on Biomedical Optics 2021, Munich, Germany, 20-24 June 2021European Conferences on Biomedical Optics 2021 (ECBO). OSA Technical Digest Optical Society of America pp. EM3B.6.
- Borri, P., Giannakopoulou, N., Zoriniants, G., Pope, I., Masia, F., Watson, P. and Langbein, W. 2019. Imaging and tracking single plasmonic nanoparticles in 3D background-free with four-wave mixing interferometry. Presented at: SPIE BIOS, San Francisco, CA, USA, 2-7 February 2019Proceedings Volume 10894, Plasmonics in Biology and Medicine XVI, Vol. 108940. Society of Photo-Optical Instrumentation Engineers (SPIE) pp. 34., (10.1117/12.2507618)
- Boorman, D., Pope, I., Langbein, W., Hood, S., Borri, P. and Watson, P. 2019. Optimisation of multimodal coherent anti-Stokes Raman scattering microscopy for the detection of isotope-labelled molecules. Presented at: SPIE BIOS, San Francisco, CA, USA, 2-7 February 2019Proceedings Volume 10890, Label-free Biomedical Imaging and Sensing (LBIS) 2019, Vol. 108900. Society of Photo-Optical Instrumentation Engineers (SPIE) pp. 4., (10.1117/12.2509280)
- Langbein, W., Harlow, D. S., Regan, D., Pope, I. and Borri, P. 2019. Heterodyne dual-polarization epi-detected CARS microscopy for chemical and topographic imaging of interfaces. Presented at: SPIE BIOS, San Francisco, California, US, 2-7 Feb 2019Label-free Biomedical Imaging and Sensing (LBIS) 2019, Vol. 10890. Bellingham, Washington: Society of Photo-optical Instrumentation Engineers (SPIE), (10.1117/12.2507636)
- Borri, P., Bradley, J., Pope, I., Langbein, W. and Swann, K. 2019. Imaging lipids in living mammalian oocytes and early embryos by coherent Raman scattering microscopy. Presented at: SPIE BIOS, San Francisco, CA, USA, 2-7 February 2019Proceedings Volume 10890, Label-free Biomedical Imaging and Sensing (LBIS) 2019, Vol. 108900. Society of Photo-Optical Instrumentation Engineers (SPIE) pp. 3., (10.1117/12.2506248)
- Pope, I. et al. 2018. Coherent Raman Scattering microscopy: technology developments and biological applications. Presented at: 20th International Conference on Transparent Optical Networks (ICTON), Bucharest, Romania, 1-5 Jul 201820th International Conference on Transparent Optical Networks (ICTON). IEEE, (10.1109/ICTON.2018.8473706)
- Borri, P., Zorinyants, G., Giannakopoulou, P., Masia, F., Pope, I. and Langbein, W. 2018. Imaging and tracking single plasmonic nanoparticles in 3D background-free with four-wave mixing interferometry. Presented at: International Conference on Transparent Optical Networks, Bucharest, Romania, 1-5 Jul 201820th International Conference on Transparent Optical Networks (ICTON). IEEE, (10.1109/ICTON.2018.8473874)
- Langbein, W. W., Rocha-Mendoza, I., Masia, F., Di Napoli, C., Pope, I., Watson, P. D. and Borri, P. 2011. Differential CARS microscopy with linearly chirped femtosecond laser pulses. Presented at: Multiphoton Microscopy in the Biomedical Sciences XI, San Francisco, CA, USA, 23-25 January 2011 Presented at Periasamy, A., König, K. and So, P. T. C. eds.Multiphoton Microscopy in the Biomedical Sciences XI, Vol. 7903. SPIE Proceedings SPIE pp. 79031I., (10.1117/12.873872)
- Flaccavento, G., Lempitsky, V., Pope, I., Barber, P., Zisserman, A., Noble, J. A. and Vojnovic, B. 2011. Learning to count cells: Applications to lens-free imaging of large fields. Presented at: Microscopic Image Analysis with Applications in Biology, Heidelberg, Germany, 2 Sept 2011.
- Njoh, K. et al. 2007. Live cell tracking on an optical biochip platform. Presented at: Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V, San Jose, CA, USA, 20 January 2007 Presented at Farkas, D. L., Leif, R. C. and Nicolau, D. V. eds.Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V. Proceedings of SPIE Vol. 6441. Bellingham, WA: SPIE pp. 64410X., (10.1117/12.698935)
- Morris, D. et al. 2007. Development of an optical biochip for the analysis of cell environment sensitivity. Presented at: Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V, San Jose, CA, USA, 20 January 2007 Presented at Farkas, D. L., Leif, R. C. and Nicolau, D. V. eds.Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues V. Proceedings of SPIE Vol. 6441. Bellingham, WA: SPIE pp. 64410V., (10.1117/12.700052)
- Matthews, D. R. et al. 2007. A fluorescence biochip with a plasmon active surface. Presented at: Plasmonics in Biology and Medicine IV, San Jose, CA, USA, 20 January 2007 Presented at Vo-Dinh, T. and Lakowicz, J. R. eds.Plasmonics in Biology and Medicine IV. Proceedings of SPIE Vol. 6450. Bellingham, WA: SPIE pp. 645006., (10.1117/12.698948)
- Thomson, J. D. et al. 2005. The role of acceptor anneal temperature on the performance of InGaN/GaN quantum well light emitting diodes. Presented at: Physics and simulation of optoelectronic devices XIII, San Jose, USA, 24-27 January 2005 Presented at Osinski, M., Henneberger, F. and Amano, H. eds.Physics and Simulation of Optoelectronic Devices XIII. Proceedings of SPIE Vol. 5722. Bellingham: SPIE pp. 425-430., (10.1117/12.591897)
Erthyglau
- Slesiona, N., Payne, L., Pope, I., Borri, P., Langbein, W. and Watson, P. 2023. Correlative extinction and single fluorophore bleaching microscopy for ligand quantification on gold nanoparticles. Advanced Materials Interfaces 10(24), article number: 2300568. (10.1002/admi.202300568)
- Pope, I., Ferreira, N. G. C., Kille, P., Langbein, W. and Borri, P. 2023. Background-free four-wave mixing microscopy of small gold nanoparticles inside a multi-cellular organ. Applied Physics Letters 122, article number: 153701. (10.1063/5.0140651)
- Pope, I. et al. 2023. Correlative light-electron microscopy using small gold nanoparticles as single probes. Light: Science & Applications 12, article number: 80. (10.1038/s41377-023-01115-4)
- Wang, Y., Pope, I., Brennan-Craddock, H., Poole, E., Langbein, W., Borri, P. and Swann, K. 2022. A primary effect of palmitic acid on mouse oocytes is the disruption of the structure of the endoplasmic reticulum. Reproduction 163(1), pp. 45-56. (10.1530/REP-21-0332)
- Boorman, D., Pope, I., Masia, F., Langbein, W., Hood, S., Borri, P. and Watson, P. 2021. Hyperspectral CARS microscopy and quantitative unsupervised analysis of deuterated and non-deuterated fatty acid storage in human cells. The Journal of Chemical Physics 155(22), article number: 224202. (10.1063/5.0065950)
- Boorman, D., Pope, I., Masia, F., Watson, P., Borri, P. and Langbein, W. 2021. Quantification of the nonlinear susceptibility of the hydrogen and deuterium stretch vibration for biomolecules in coherent Raman microspectroscopy. Journal of Raman Spectroscopy 52(9), pp. 1540-1551. (10.1002/jrs.6164)
- Pope, I. et al. 2021. Identifying subpopulations in multicellular systems by quantitative chemical imaging using label-free hyperspectral CARS microscopy. Analyst 146(7), pp. 2277-2291. (10.1039/D0AN02381G)
- Nahmad-Rohen, A., Regan, D., Masia, F., McPhee, C., Pope, I., Langbein, W. and Borri, P. 2020. Quantitative label-free imaging of lipid domains in single bilayers by hyperspectral coherent Raman scattering. Analytical Chemistry 92(21) (10.1021/acs.analchem.0c03179)
- Giannakopoulou, N. et al. 2020. Four-wave-mixing microscopy reveals non-colocalisation between gold nanoparticles and fluorophore conjugates inside cells. Nanoscale 12(7), pp. 4622-4635. (10.1039/C9NR08512B)
- Lloyd, D. et al. 2020. Functional imaging of a model unicell: Spironucleus vortens as an anaerobic but aerotolerant flagellated protist. Advances in Microbial Physiology 76, pp. 41-79. (10.1016/bs.ampbs.2020.01.002)
- Bradley, J., Pope, I., Wang, Y., Langbein, W., Borri, P. and Swann, K. 2019. Dynamic label-free imaging of lipid droplets and their link to fatty acid and pyruvate oxidation in mouse eggs. Journal of Cell Science 132(13), article number: jcs228999. (10.1242/jcs.228999)
- Langbein, W., Regan, D., Pope, I. and Borri, P. 2018. Heterodyne dual-polarization epi-detected CARS microscopy for chemical and topographic imaging of interfaces. APL Photonics 3, article number: 92402. (10.1063/1.5027256)
- Masia, F., Pope, I., Watson, P., Langbein, W. and Borri, P. 2018. Bessel-beam hyperspectral CARS microscopy with sparse sampling: enabling high-content high-throughput label-free quantitative chemical imaging. Analytical Chemistry 90(6), pp. 3775-3785. (10.1021/acs.analchem.7b04039)
- Bradley, J., Pope, I., Masia, F., Sanusi, R., Langbein, W., Swann, K. and Borri, P. 2016. Quantitative imaging of lipids in live mouse oocytes and early embryos using CARS microscopy. Development 143, pp. 2238-2247. (10.1242/dev.129908)
- Di Napoli, C., Pope, I., Masia, F., Langbein, W., Watson, P. and Borri, P. 2016. Quantitative spatiotemporal chemical profiling of individual lipid droplets by hyperspectral CARS microscopy in living human adipose-derived stem cells. Analytical Chemistry 88(7), pp. 3677-3685. (10.1021/acs.analchem.5b04468)
- Pope, I. et al. 2014. Coherent anti-Stokes Raman scattering microscopy of single nanodiamonds. Nature Nanotechnology 9(11), pp. 940-946. (10.1038/nnano.2014.210)
- Di Napoli, C., Pope, I., Masia, F., Watson, P. D., Langbein, W. W. and Borri, P. 2014. Hyperspectral and differential CARS microscopy for quantitative chemical imaging in human adipocytes. Biomedical Optics Express 5(5), pp. 1378-1390. (10.1364/BOE.5.001378)
- Di Napoli, C., Masia, F., Pope, I., Otto, C., Langbein, W. W. and Borri, P. 2014. Chemically-specific dual/differential CARS micro-spectroscopy of saturated and unsaturated lipid droplets. Journal of Biophotonics 7(1-2), pp. 68-76. (10.1002/jbio.201200197)
- Pope, I., Langbein, W. W., Watson, P. D. and Borri, P. 2013. Simultaneous hyperspectral differential-CARS, TPF and SHG microscopy with a single 5 fs Ti:Sa laser. Optics Express 21(6), pp. 7096-7106. (10.1364/OE.21.007096)
- Pope, I., Barber, P. R., Horn, S., Ainsbury, E., Rothkamm, K. and Vojnovic, B. 2011. A portable microfluidic fluorescence spectrometer device for γ-H2AX-based biological dosimetry. Radiation Measurements 46(9), pp. 907-911. (10.1016/j.radmeas.2011.02.004)
- Pope, I., Langbein, W. W., Borri, P. and Watson, P. D. 2011. CARS imaging for high throughput microscopy. Biotech International 23(Apr/Ma), pp. 11-13.
- Thomson, J. D. et al. 2006. The influence of acceptor anneal temperature on the performance of InGaN/GaN quantum well light-emitting diodes. Journal of Applied Physics 99(2), article number: 24507. (10.1063/1.2165405)
- Brown, I. H. et al. 2005. Determination of the Piezoelectric Field in InGaN Quantum Wells. Applied Physics Letters 86, article number: 131108. (10.1063/1.1896446)
- Pope, I., Smowton, P. M., Blood, P., Thomson, J. D., Kappers, M. J. and Humphreys, C. J. 2003. Carrier leakage in InGaN quantum well light-emitting diodes emitting at 480 nm. Applied Physics Letters 82(17), pp. 2755-2757. (10.1063/1.1570515)
Gosodiad
- Pope, I. 2004. The characterisation of InGaN/GaN quantum well light emitting diodes. PhD Thesis, Cardiff University.
Patentau
- Vojnovic, B., Barber, P., Pope, I., Smith, P. J. and Errington, R. J. 2008. Detecting objects. WO/2008/090330 [Patent].
Research
Coherent Anti-Stokes Raman Scattering (CARS)
In CARS, two near infra-red laser beams of different frequencies (called "pump" (ω1) and "Stokes" (ω2)) are used to drive the molecular vibrations at the frequency ωD = ω1 - ω2 (where, ω1>ω2). By adjusting the frequency of the Stokes beam the driving frequency can be tuned to specific vibrational resonance (ωD = Ω). A third beam (ω3) is then used to generate the emission of a photon (ω4) at the frequency ω4 = ω1 - ω2 + ω3, which is detected. Usually ω1 and ω3 are chosen to be degenerate therefore, ω4 = 2ω1 - ω2 = ω1 + Ω. Hence ω4 is the anti-Stokes Raman frequency relative to ω1 . Since all bonds within the focal volume with a vibrational resonance equal to Ω are drive coherently, the anti-Stokes Raman emission constructively interferes with itself resulting in a non-linear increase in signal. Since the CARS signal scales non-linearly with the number of bonds within the focal volume, CARS is ideally suited to imaging compounds with a large number of similar bonds within their structure, such as lipids, which contain a large number of C-H bonds within their acyl chains.
CARS energy level diagram. E1 and E0 are electronic levels, with E1 - E0 typically UV for molecules studied with CARS. Ω is the energy separation between vibrational modes, typically mid-infrared. Dashed lines represent virtual energy states.
http://dx.doi.org/10.1016/B978-0-12-391857-4.00014-8
Our Single Source CARS/TPF/SHG microscope
We have developed a multimodal multiphoton laser-scanning microscope for cell imaging featuring simultaneous acquisition of D-CARS, TPF and SHG using a single 5 fs Ti:Sa broadband (660-970 nm) laser. The spectral and temporal pulse requirements of these modalities were optimized independently by splitting the laser spectrum into three parts: TPF/SHG excitation (> 900 nm), CARS Pump excitation (< 730 nm), and CARS Stokes excitation (730-900 nm).
CARS Beam Optimisation
After splitting the initial broadband laser pulse up into the pump and Stokes beams each generated femtosecond pulse is still much broader than the typical Raman linewidths (i.e., the duration of pump and Stokes pulses are much shorter than the vibrational coherence time which are on the order of a picosecond). However, the vibrational excitation in CARS is governed by the interference of the pump and Stokes fields; therefore, the spectral resolution is not determined by these individual spectra, but by the spectrum of their temporal interference. Hence by carefully shaping the pump and Stokes pulses in time, it is possible to drive a narrow vibrational frequency range, even though the individual pulses are spectrally broad. We recently demonstrated a simple, highly efficient, alignment insensitive, and cost-effective method which utilizes glass elements of known dispersion (Langbein et al., 2009a; Rocha-Mendoza et al., 2008) through a process known as spectral focusing or chirp.
Spectral Focusing
The refractive index (n) of a dispersive medium is not constant but varies with wavelength, n(lambda) (and thus frequency). This means that when a short pulse travels through a dispersive medium, its different wavelength (or frequency) components travel at different speeds Typically, the longer wavelengths travel faster and thus emerge from the medium first. This stretches out the pulse from its initial width. The key point for spectrally selective CARS is to introduce the same chirp parameter in pump and Stokes pulses so that the instantaneous frequency difference (IFD) between the two pulses remains constant. The molecular vibrations are then driven at the beat frequency (w1-w2) which will be centred at the IFD. Using glass dispersion to chirp the beams we achieve a CARS spectral resolution of 10 cm−1. Tuning to different Raman frequencies is achieved by simply by adjusting the arrival time of the pump beam relative to the Stokes. Using this method we are able to acquire CARS images over the spectral range 1200-3800 cm−1.
Differential CARS (D-CARS)
D-CARS is implemented with few passive optical elements and enables simultaneous excitation and detection of two vibrational frequencies with a separation adjustable from 20 cm−1 to 150 cm−1 for selective chemical contrast and background suppression.
(a) Broadband sub-10 fs lasers produce a broad range of wavelengths, that can be separated (b) into pump and Stokes beams through the use of a dichroic mirror (DM). (c) Illustration demonstrating that the IFD between two equally chirped pulses is constant with time. (e) Illustration demonstrating how varying the overlap of the two pulses changes the IFD, thus allowing spectral tuning via simple delay scanning. t, time; ω, frequency.
TPF/SHG Beam Optimisation
A prism pulse compressor in the TPF/SHG excitation is used to achieve Fourier limited 30 fs pulses at the sample for optimum TPF and SHG.
A sketch of the microscope set-up. M: mirror; DM: dichroic mirror; SF57: glass blocks; R: reflecting prism; λ /2: half-wave plate; BE: beam expander; PBS: polarizing beam splitter; F: filter. The side view of the optics between by the two indicated arrows shows the beam height difference. Graph: typical spectra of the laser, pump, Stokes and TPE beams.
Mouse tail tissue section fluorescently labeled with FITC-Concanavalin A. Top Left: Stitched epi-fluorescence image, scale bar = 300μm. Bottom Left: Single epi-fluorecence image cropped to 150×150μm (the boxed region shown on the Top Leftimage). Far Right: False-colour image generated from simultaneously acquired TPF, CARS and SHG images. CARS from subcutaneous lipid deposits at 2850 cm-1 (red) and SHG from collagen (blue), scale bar = 20μm. 150×150μm, 501×501 pixels, 0.01 ms pixel dwell time (2.5 s total image acquisition).
http://dx.doi.org/10.1364/OE.21.007096
Hyperspectral Imaging
The images below were generated by taking a hyperspectral CARS stack of images at different molecular vibrations. Through the analysis of this image stack a spectral profile for each pixel in the image may be generated. Further analysis of these profiles allows a false-colour chemical map may be generated. (red: Lipids, blue: DNA, green: protein)
A human metastatic colorectal carcinoma imaged live using our CARS microscopy. The 3D culture system used to grow the carcinoma spheres aims to reproduce the architecture of the cancer, allowing us to employ CARS microscopy to investigate the composition of these spheres outside the patient's body.
A mouse intestinal organoid, images live using our CARS microscope. The 3D culture system recapitulates the architecture of the small intestine with crypt and villi structures, comprising both stem cells and differentiated cells.
Arnica Karuna, Andrew Hollins, Iestyn Pope, Anika Offergeld , Francesco Masia, Wolfgang Langbein, Trevor Dale, Paola Borri
Nanodiamonds
Nanoparticles are attracting enormous attention for biomedical applications as optical labels, drug delivery vehicles and contrast agents in vivo. Nanoparticles made of various types of organic and inorganic materials have been investigated, and recently diamond has emerged as one of the best material choices due to its bio-compatibility and unique structural, chemical, mechanical, and optical properties. So far, diamond nanoparticles (nanodiamonds) have been visualised optically mainly via the existence of fluorescing 'defects' in the crystal lattice. However, the production of these defects is costly and not very well controlled hence limiting their use. Moreover, these defects might irreversibly change their fluorescence properties upon optical illumination and when close to the surface of a nanodiamond they might become unstable. We have shown that single non-fluorescing nanodiamonds exhibit a strong CARS signal at the sp3 vibrational resonance of diamond. Using our in-house developed CARS microscope we have measured the CARS signal strength on a series of single nanodiamonds of different sizes. The nanodiamond size was accurately determined by means of electron microscopy and correlative quantitative optical contrast methods developed in-house. In this way, we were able to quantify the relationship between the CARS signal strength and nanoparticle size. The calibrated CARS signal in turn enables us to analyse the number and size of nanodiamonds internalized in living cells in situ. Owing to the high bio-compatibility of nanodiamonds, this imaging modality opens the exciting prospect of following complex cellular trafficking pathways quantitatively.
http://dx.doi.org/10.1038/nnano.2014.210
A 3D CARS image of a HeLa cell that has taken up nanodiamonds.  The image consists as an overlay of the CARS signal at the nanodiamond resonance (1332 cm-1) and the CARS signal from intracellular structures (2900 cm-1) where the relative intensities have been adjusted to enhance the cellular contrast for visualisation purposes. A false colour map has then been used where the nanodiamonds can be seen as the bright orange spots within the magenta cell.
Lipids
We have demonstrated the applicability of CARS micro-spectroscopy for quantitative chemical imaging of saturated and unsaturated lipids in human stem cell derived adipocytes. Comparing dual-frequency/differential CARS (D-CARS), which enables rapid imaging and simple data analysis, with broadband hyperspectral CARS microscopy analysed using an unsupervised phase-retrieval and factorization method recently developed within our group for quantitative chemical image analysis.
Through a ratiometric analysis, both D-CARS and phase-retrieved hyperspectral CARS determine the concentration of unsaturated lipids with comparable accuracy in the fingerprint region, while in the CH stretch region D-CARS provides only a qualitative contrast owing to its non-linear behaviour. When analysing hyperspectral CARS images using the blind factorization into susceptibilities and concentrations of chemical components recently demonstrated by us, we are able to determine vol:vol concentrations of different lipid components and spatially resolve inhomogeneities in lipid composition with superior accuracy compared to state-of-the art ratiometric methods.
Biography
I did my undergraduate Master of Physics degree at Cardiff University, graduating in 2000. Followed by Ph.D (2000-2004) in the area of optoelectronics on "The characterisation of InGaN/GaN quantum well light emitting diodes" under the supervision of Prof P. Blood and Prof P. Smowton.
From 2005 to 2007 I worked as a research scientist in the Advanced Technology and Development Group at the Gray Cancer Institute. Here I branched over into the area of Biophysics, developing microscopy-related and other novel instruments for biological research. In 2009 at The Engineer Technology and Innovation awards, presented at The Royal Society, one of these devices "CyMap" won the Medical and Healthcare award and the overall Grand Prix award.
Following the groups relocation from the Cray Cancer Institute to the Gray Institute for Radiation Oncology and Biology at the University of Oxford I worked on the development of a portable biological dosimeter for the rapid classification of casualties following a radiological incident (2007 to 2010). The prototype device employed spectral un-mixing to determine exposure doses from white blood cells stained with fluorescent antibodies conjugated to the gamma-H2AX histone. Winning the poster prize for best young researcher at the EPRBioDose conference held in Mandelleu-LaNapoule (France), 2010.
In 2010 I returned to Cardiff University to join the BioPhotonics & Quantum Optoelctronics Group as a research associate to work on the development of a new generation single source laser-scanning multiphoton microscope. The microscope capable of simultaneously imaging with D-CARS, TPF and SHG is know complete and forms an integral part of the groups research activities. My current research activities include the study of lipid droplet dynamics in biological samples such as HeLa cell, HeG2 cells and oocytes; The development of the CARS microscope to move towards high-throughput hyperspectral imaging; The use of nanodiamonds as optical labels and contrast agents in live cells; and the use of resonant Four-Wave Mixing (FWM) to image gold and silver nanoparticles inside cells.
Honours and awards
The Engineer Awards (4 December 2009)
- Winner of the Medical/Health Care category, with CyMap.
- Winner of the Grand Prix: The Engineer Special Award, with CyMap.
EPR Biodose (10-14th October 2010)
- Poster Prize for best young scientist.
Bio-Nano-Photonics (13-14th September 2011)
- Poster Prize
Contact Details
+44 29208 79037
Sir Martin Evans Building, Room Cardiff School of Biosciences, The Sir Martin Evans Building, Museum Avenue, Cardiff, CF10 3AX, Museum Avenue, Cardiff, CF10 3AX