Pingyong Xu

Biography

1992-1996 Central China Normal University, B.S. in Chemical Education, Wuhan, China 1996-2000 Central China Normal University, M.S. in Organic Chemistry, Wuhan, China 2000-2004 Huazhong University of Science and Technology, Ph.D. in Engineering of Biomedicine, Wuhan, China 2004 - 2006 Postdoctoral Fellow, Assistant Professor,Institute of Biophysics, Chinese Academy of Sciences, Beijing, China 2006 - 2010 Associate Professor, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China 2010 - Professor, Institute of Biophysics, Chinese Academy of Sciences, Beijing, China

Research Interest

Our studies focus on two aspects: one combines spectroscopy, biophysical microscopy techniques, and protein design and engineering for the development of novel optical imaging tools, especially photocontrollable fluorescent proteins and functional chemical dyes. In addition, our research uses the probes we develop in SR techniques to study important biological processes such as autophagosome formation. Using fluorescence microscopy, information about the spatial organization of specific target proteins can be accurately provided at the molecular level by labeling proteins with fluorescent proteins. However, conventional optical microscopy is limited by diffraction to imaging on a coarser scale (~250 nm) by two orders of magnitude. Recently, several SR imaging techniques, such as single molecule localization-based microscopy (PALM/FPALM/STORM) and structured illumination-based techniques such as nonlinear structured illumination microscopy (NL-SIM), were developed for the study of cellular ultrastructure at diffraction-unlimited resolutions. However, many efforts are still needed to improve the spatial and temporal resolutions and labeling technology, especially in living cells. One of the major limits is the absence of appropriate fluorescent probes with specific photochemical properties. Different SR techniques use different fluorescent probes with optimized properties. Live-cell SR imaging is also very challenging because it requires fluorescent proteins to be very stable and bright and have a high contrast ratio. We mainly focus on developing novel fluorescent probes, especially fluorescent proteins that show priority to dyes in living cells, for diffraction-unlimited optical microscopy. We hope to further improve the spatial-temporal resolution using multiple fluorescent labels with higher photon numbers and contrast ratios. Although many current SR techniques have been successfully demonstrated to image cellular dynamics, applications have been rather limited and appear challenging. Live-cell STED/reversible saturable optical fluorescence transition (RESOLFT) and SIM/nonlinear SIM require sophisticated and expensive optical setups and professional expertise for accurate optical alignment. Live-cell PALM/STORM uses a less complicated setup; however, a sCMOS camera, whose pixel-dependent noise should be pre-characterized and calibrated before use, is required for extremely high acquisition speeds over tens of thousands of frames. Recently, wide-field based SR microscopies have been developed to improve temporal resolution using much fewer time-lapse images (hundreds to thousands) than PALM/STORM. One of them, a Bayesian analysis of blinking and bleaching (3B), offers enormous potential to resolve ultrastructure and fast cellular dynamics beyond the diffraction limit in living cells. Despite the potential, the 3B analysis is impractical when imaging the nanoscale dynamics of large fields of view in live cells over long time periods, as the calculation is extremely time-consuming and/or consumes large amounts of web resources. Another major problem for 3B imaging is the artificial thinning and thickening of structures both in simulated images and experimental data. Our goal is to develop simple but useful SR techniques with high spatial-temporal resolution. We are developing novel algorithms and imaging techniques based on a simple TIRFM system to make them useful SR imaging tools in common labs for both fixed and living cells. Autophagy is a complex, multi-step, and biologically important pathway mediated by autophagosomes and autolysosomes. Accurately dissecting and detecting different stages of autophagy is important to elucidate the molecular mechanism and thereby facilitate the discovery of pharmaceutical molecules. We have been developing fluorescent dyes to dynamically monitor intracellular organelle localization and functional probes for autophagosome formation.In the future we will use SR techniques to imaging the dynamic formation of intracellular organelles such as autophagosome and ER.