Robert M. Clegg
Professor of Physics, OF Bioengineering, and of Biophysics
Robert MacDonald Clegg received his doctorate in physical chemistry in 1974 from Cornell University. His dissertation is entitled "Relaxation Kinetics Applying Repetitive Pressure Perturbations," was supervised by Professor E. L. Elson. Following graduation, Professor Clegg moved to Göttingen, Germany, where he started work as a postdoctoral research associate in the Max Planck Institute for Biophysical Chemistry. In 1976, he was promoted to Senior Staff Research Associate in the Department of Molecular Biology. Dr. Clegg remained at the Max Planck Institute until he accepted a position as Professor of Physics at the University of Illinois at Urbana-Champaign in 1998. Presently, Dr. Clegg is a Professor in the Departments of Physics and Bioengineering and is on the faculty of the Biophysics Program at the University of Illinois.
Research Interests: Professor Clegg's research focus is in nucleic acid (DNA/RNA) structures and conformational changes (branches, junctions, bulges); protein-nucleic acid interaction: histones, polymerases, structure-recognizing binding proteins. Interaction of small molecules (dyes) with DNA: intercalation, groove binding, enzyme inhibition; transcription: catalysis of RNA synthesis, physical studies, enzyme assembly-activity; lipid and small protein diffusion processes in membranes; high pressure (HP) studies of multisubunit molecular assemblies, viruses and inactivation of enzymes; HP electrophoresis; HP spectroscopy; HP enzymology; stopped flow, microsecond fast flow mixing; relaxation kinetics (pressure- and temperature-jump; application of fluorescence in biophysics; energy transfer for structural determinations; fluorescence to other physical techniques; nanosecond lifetime studies; delayed luminescence imaging microscopy (DLIM) -phosphorescence and delayed luminescence; fluorescence lifetime-resolved imaging microscopy (FLIM) in nanosecond times; fluorescence diagnosis of lung carcinomas using lifetime-resolved imaging endoscopy.
Description of Current Research
Development of Lifetime-resolved Advanced Spectroscopy Fluorescence Microscopes
The scope of new applications in optical microscopy has increased spectacularly in the biological sciences in recent years, primarily because of the simultaneous advances in technology and in chemical and biological methodology. Techniques such as real-time video visualization and rapid digital image acquisition permit sensitive observation of dynamic processes in vivo, and
various experimental innovations, such as fluorescence correlation spectroscopy (FCS), fluorescence recovery after photobleaching (FRAP), single molecule observation, etc., extend the conventional use of microscopy and broaden its applications into diverse fields. The concomitant development of extrinsic probes and their attachment procedures, and the rapid and seemingly unending
growth of biotechnological techniques, have provided powerful biological tools which expand considerably the applications of fluorescence microscopy beyond the usual modus operandi of localizing and estimating the relative amount of specific structures and molecules. This expansion of optical microscopy into new fields of biology, and the confluence of instrumentation technical developments with
biotechnology has given fluorescence microscopy the well earned reputation as one of the major means for acquiring vital information on the functioning of biological systems.
Developments that have considerably benefited microscopy have been made in fields of microscope technology (confocal capabilities, scanning stages, full field microscopes with increased sensitivity and improved optical performance, etc.), optics (e.g. multi-photonic excitation modalities), lasers (e.g., tunable, solid state, femtosecond lasers), computers, chemistry of indicators, immunology, genetics, and molecular biology. To a great extent, the crucial role of fluorescence imaging has been due to its superb sensitivity and specificity. The major—and in most cases the sole—fluorescence parameter measured quantitatively in microscopy is the intensity of the emitted light integrated over a selected wavelength range. We are proposing the introduction of new spectroscopic and physical measurements into the fluorescence microscope which will extend appreciably the capabilities of fluorescence microscopy. The major objective is an extension of the capabilities of fluorescence microscopy, not an incremental improvement in currently available imaging technologies. New physical information will be obtained by determining directly the dynamic characteristics of the fluorescence emission (lifetime), of the fluorescence polarization and of the dispersion of the emission spectrum.
The incorporation of these capabilities into a practical, conveniently operated fluorescence microscope with three-dimensional imaging capabilities entails a major development and engineering project. New perspectives for experiments are opened up by bringing more quantitative physical measurements into the microscope setting. Applications of these new microscopic measurements to several currently vital biological research areas, such as receptor function, protein/lipid interactions, membrane domains, chromatin structure and apoptosis.
The new spectroscopic features we wish to incorporate into the fluorescence microscope have been proven to be extremely useful in macroscopic fluorescence measurements. At present, these spectroscopic capabilities are available only on optical microscopes in a few special laboratories, such as ours. Fluorescence measurements can contribute much more information than just the location of a compound, or an approximation of relative concentrations. Lifetime-resolved fluorescence techniques allow the physical characteristics of a chromophore’s immediate neighborhood to be probed. By developing and consolidating these advanced spectroscopy microscope measurements, and making these methods more accessible for routine measurements, they will become appreciated, and applied, by those practicing biology and microscopy, opening up new opportunities for biological investigations.
Selected Publications
Redford, GI, Majumdar, ZK, Sutin, JDB, and Clegg, RM. Properties of microfluidic turbulent mixing revealed by fluorescence lifetime imaging. J. Chem. Phys. 123, 224504-1-6 (2005).
Eckhoff, DA, et al. Optical characterization of ultrasmall Si nanoparticles prepared through electrochemical dispersion of bulk Si. J. Phys. Chem. B 109, 19786-19797 (2005).
Redford, GI and Clegg, RM. Polar plot representation for frequency-domain analysis of fluorescence lifetimes. J. Fluoresc. 15, 805-815 (2005).
Rao, S, et al. Excited states of tetrahedral single-core Si29 nanoparticles. Phys. Rev. B 69, 205319-1-7 (2004).
Clegg, RM. The vital contributions of Perrin and Forster. Biophotonics Intl. 11, 42-45 (2004).