Brown University School of Engineering

School of Engineering & Department of Physics Joint “Electronics and Photonics Seminar” & “Condensed Matter and Biological Physics Seminar” Peering into Cells One Molecule at a Time: Single-molecule and plasmon-enhanced fluorescence for super-resolution i

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Thursday, September 27, 2012 4:00pm - 5:00pm

ost: Prof. Domenico Pacifici, School of Engineering Sponsored by ADVANCE at Brown under NSF Grant No. 0548311. School of Engineering & Department of Physics Joint “Electronics and Photonics Seminar” & “Condensed Matter and Biological Physics Seminar” Peering into Cells One Molecule at a Time: Single-molecule and plasmon-enhanced fluorescence for super-resolution imaging of living bacteria Prof. Julie Biteen Assistant Professor of Chemistry and Biophysics University of Michigan, Ann Arbor, MI Abstract: Single-molecule fluorescence brings the resolution of optical microscopy down to the nanometer scale, allowing us to unlock the mysteries of how biomolecules work together to achieve the complexity that is a cell. This high-resolution, non-destructive method for examining subcellular events has opened up an exciting new frontier: the study of macromolecular localization and dynamics in living cells. We have developed methods for single-molecule investigations of live bacterial cells, and have used these techniques to investigate three important prokaryotic systems: membrane-bound transcription activation in Vibrio cholerae, carbohydrate catabolism in Bacteroides thetaiotaomicron, and DNA mismatch repair in Bacillus subtilis. Each system presents unique challenges, and we will discuss the important methods developed for each system, in particular, a comparison of membrane-bound and soluble proteins, extensions to two-color and 3D imaging, and adaptations for studying live anaerobic cells. Furthermore, we use the plasmon modes of bio-compatible metal nanoparticles to enhance the emissivity of single-molecule fluorophores. The resolution of single-molecule imaging in cells is generally limited to 20-40 nm, far worse than the 1.5-nm localization accuracies which have been attained in vitro. We therefore use plasmonics to improve the brightness and stability of single-molecule probes, and in particular fluorescent proteins, which are widely used for bioimaging. We find that gold-coupled fluorophores demonstrate brighter, longer-lived emission, yielding an overall enhancement in total photons detected. Ultimately, this results in increased localization accuracy for single-molecule imaging. Furthermore, since fluorescence intensity is proportional to local electromagnetic field intensity, these changes in decay intensity and rate serve as a nm-scale read-out of the field intensity. Our work indicates that plasmonic substrates are uniquely advantageous for super-resolution imaging, and that plasmon-enhanced imaging is a promising technique for improving live cell single-molecule microscopy. Biosketch: Julie Biteen has been an Assistant Professor of Chemistry with courtesy appointments in Biophysics and Applied Physics at the University of Michigan since January 2010. Dr. Biteen did an A.B. in Chemistry at Princeton University before moving to California Institute of Technology, where she studied plasmon-enhanced emission from silicon quantum dots in the labs of Harry Atwater and Nathan Lewis. She received a Masters in Applied Physics and a Ph.D. in Chemistry at Caltech, before going on to a postdoc in the lab of W. E. Moerner at Stanford University studying structural proteins in Caulobacter crescentus cells with single-molecule imaging. Dr. Biteen is the recipient of a Burroughs Wellcome Career Award at the Scientific Interface and a PicoQuant Young Investigator Award, and has been named a University of Michigan Biological Scholar. Research in the Biteen Lab focuses on single-molecule super-resolution imaging of biomolecular localization, cooperativity, and dynamics in bacteria and on using plasmonic nanoparticles to enhance emission and therefore single-molecule resolution in these bio-imaging applications. Thursday, September 27th 4:00