77 76 Principal Scientist Profiles Christian Eggeling Christian Eggeling Principal Scientist Profiles PROFESSOR FOR SUPERRESOLUTION MICROSCOPY AND INSTITUTE DIRECTOR, INSTITUTE OF APPLIED OPTICS AND BIOPHYSICS AND PROFESSOR OF MOLECULAR IMMUNOLOGY Dr. Eggeling holds a PhD in Physics from the University of Göttingen, was then a research scientist at the biotech company Evotec, Hamburg, before joining the department of Professor Stefan Hell (2014 Nobel Laureate in Chemistry) at the Max-Planck-Institute for Biophysical Chemistry in Göttingen. In 2012, he started as a principal investigator in the MRC Human Immunology Unit and as the scientific director of the newly established Wolfson Imaging Centre Oxford at the Weatherall Institute of Molecular Medicine, University of Oxford, and was appointed Professor of Molecular Immunology in 2014, positions which he still holds today. In 2017, he started as a Professor of Superresolution Microscopy and director of the Institute of Applied Optics and Biophysics (IAOB) at the Friedrich Schiller University Jena, and as the Head of the Department of Biophysical Imaging at the Leibniz IPHT Jena. CHRISTIAN EGGELING RESEARCH AREAS The research group of Christian Eggeling is focused on the development of advanced microscopy for the investigation of molecular organization and dynamics in cells, especially on the cellular plasma membrane. Highlights are the optimization of superresolution STED microscopy and its combination with single-molecule fluorescence spectroscopy tools such as fluorescence correlation spectroscopy (FCS), use of adaptive optics for deep-tissue investigations, advancements in singleparticle tracking (using fluorescence, interferometric scattering (iSCAT) and novel superresolution MINFLUX microscopy), the detailed investigation of lipid membrane heterogeneity such as lipid rafts, and biological applications of all of these tools for investigations of multiple biomedical issues such as within the Excellence Cluster “Balance of the Microverse”, the Collaborative Research Center 1278 PolyTarget, infection diagnostics or immunology. Further, fully serviced user microscope facilities have been set up and are being optimized. TEACHING FIELDS Main teaching activities include bachelor biophysics lectures and exercises, master applied laser technology lectures and exercises as well as support of physics teaching practical. Further, student assistants, master and bachelor as well as PhD students are welcome and supported through various research projects. RESEARCH METHODS The Eggeling group is specialized on advanced fluorescence microscopy techniques, especially superresolution STED microscopy in combination with fluorescence correlation spectroscopy (STED-FCS), and has access to multiple microscopes including confocal, wide-field/TIRF/MINIFLUX superresolution, structured illumination and STED microscopes, but is also using complementary approaches such as single-particle tracking and intererometric Scattering (iSCAT) microscopy. In addition to that, the group has access to biochemical wet labs, cell culture and optical labs (up to biosafety level 2). RECENT RESEARCH RESULTS Recent research includes the use of artificial intelligence algorithms for an optimized fluorescence microscopy/ spectroscopy analysis [1], advanced microscopy of molecular interactions involved in virus infection [2-4], use of adaptive optics for optimized inner-cellular and tissue observations [5, 6], and the biophysical characterization of immune responses [7, 8]. DEVELOPMENT OF STED-FCS The Eggeling group has developed and in applications further advanced the combination of superresolution STED microscopy with fluorescence correlation spectroscopy (STED-FCS) to in more detail investigate molecular interactions, especially in the cellular plasma membrane, elucidating long-standing problems such as lipid membrane heterogeneity, e.g. lipid rafts. See for example Eggeling et al., Nature 457, 1159 (2009) or Sezgin et al., Nature Protoc. 14, 1054 (2019). Figure: a, Representative confocal and STED microscopy images of immobilized 40-nm fluorescent beads. The confocal and STED laser beams are well aligned when these images are perfectly centered. b, A representative intensity line profile for a single isolated bead from the images in a (green: confocal; magenta: STED), confirming the optimized alignment. These profiles can be used to obtain the diameters (fullwidth-half-maximum [FWHM] values) of the imaged spots (d). c, Representative confocal (left, green) and STED (right panels, magenta) images of fluorescent beads for different time delays between the excitation and STED laser pulses; perfect timing is at 0 ns. Figure: STED-FCS measurements of the diffusion of the viral protein ENV on HIV-1 virus particles. a Live confocal imaging was used to locate individual HIV-1 virus particles in a 2 micron × 2 micron imaging window using the signal from a labeled viral protein (green) as a guide and to align them with the position of a beamscanning line (red). Scale bar: 200 nm. b, c Representative signal for each point along the scanned line over time (intensity carpet) for the viral protein (green) in confocal mode and the Env surface protein (orange) in superresolution STED mode on wild-type mature HIV-1 particles. Image x- and y-axis correspond to the position on the scan line and signal intensity at each time point, respectively. Scale bars: x-axis (white) = 200 nm, y-axis (grey) = 4.4 ms. d FCS correlation curve for each point of the scanned line (correlation carpet) generated from signal intensity carpet shown in c. Image x- and y-axis correspond to correlation time tau and the position on the scan line, respectively. Colour code corresponds to the normalised FCS autocorrelation curves G(tau) at each position on the scan line. e Representative normalised autocorrelation curves of Env diffusion (grey and black lines) obtained from individual positions on the scan line within correlation carpets. Autocorrelation curves were fitted (coloured lines) for mature (red), immature (blue) and completely fixed HIV-1 particles (purple) using generic two-dimensional diffusion model. Greyed out area corresponds to the photobleaching-only portion of the correlation data. The data highlights faster diffusion on mature compared to immature viruses, with fully fixed viruses showing the correlation curve decay due to pure photobleaching. [1] Waithe et al., J. Cell Biol. 219, e201903166 (2020). [2] Favard et al., Science Adv. 5, eaaw8651 (2019). [3] Chojnacki et al., Nature. Commun. 8, 545 (2017). [4] Carravilla et al., Nature. Commun. 10, 78 (2019). [5] Barbotin et al., Opt. Express 27, 23378 (2019). [6] Barbotin et al., ACS Photonics 7, 1742 (2020). [7] Fritzsche et al., Sci. Adv. 3, e1603032 (2017). [8] Santos et al., Nat. Immunol. 19, 203 (2019). Contact: Phone: + 49 3641 9-47670 Email: christian.eggeling@uni-jena.de
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