While technology such as x-ray imaging, magnetic resonance imaging and electron microscopy represent milestones that have enabled us to study physiology in unprecedented detail, our knowledge is still far from being comprehensive. The structural complexity of cells and organisms is further complicated by processes involving an enormous range of spatial and temporal scales. The challenge is to tease these processes apart bit-by-bit. Optical methods, summarized under the term biophotonics, enrich the toolbox helping us with this tedious but important task.
The first part of this talk will be focused on how to capture dynamic processes on the nanoscale using super-resolution techniques paired with fluorescence fluctuation spectroscopy. Biomedical applications include the characterization of the movement of natural killer cell receptors to help understand how their response towards tumor cells is modulated. The second part will be centered on the application of light sheet microscopy to image transient events in single cells, in tissues as well as in entire organisms. For this purpose, we devised a flexible multipurpose platform for light sheet microscopy, the sideSPIM. A wide range of sample volumes (10-1,000 µl) and geometries can be accommodated and different imaging conditions can be adopted quickly. This was shown for a large variety of applications such as capturing the microcirculation of erythrocytes in a zebrafish embryo, studying the growth of bacteria biofilms under flow using a fluidic device, and the quantification of the diffusion of proteins in mammalian cells and plant roots by means of fluorescence fluctuation based image mean square displacement (iMSD) and two-dimensional pair cross-correlation function (2D-pCF) analysis. We further evaluated the potential of the sideSPIM system for high throughput screening applications, especially regarding tumor tissue samples. By adding capability for spectral and fluorescence lifetime detection, we are seeking to gain as much information about the sample as possible. The future challenge is to extend the big data concept by smart data acquisition in order to access as many spatial and temporal scales as possible.
Per Niklas Hedde, Ph.D., is a postdoctoral researcher at the University of California, Irvine where he develops light sheet microscopy, camera-based fluorescence fluctuation spectroscopy techniques and particle counting devices for medical diagnosis and antibody discovery. He studied physics at the University of Ulm, Germany, with a master thesis project on ultrafast analysis of super-resolution microscopy data. He completed his Ph.D. in physics at the Karlsruhe Institute of Technology where he built an instrument for stimulated emission depletion based raster image correlation spectroscopy to study the dynamics of cell membrane receptors and applied dual color photoactivation localization microscopy to image desmin protein mutations related to heart disease. For his thesis work he received the Karlsruhe Institute of Technology Award for Outstanding Doctoral Research Work in the Area of Applied Life Sciences 2014 and the Gregorio Weber International Prize 2014. He then accepted a postdoctoral position at the Laboratory for Fluorescence Dynamics at UC Irvine to broaden his skills including fluorescence lifetime, spectral and polarization imaging. During this time, he also visited and collaborated with the Karolinska Institute in Stockholm, Sweden to learn about natural killer cells, their value for cancer immune therapy and to establish fluctuation spectroscopy methods at the KI Department of Microbiology, Tumor and Cell Biology. He has published 25 peer-reviewed journal articles and is member of the Biophysical Society (US and Germany), the American Association for Cancer Research and the UC Irvine Center for Complex Biological Systems.