An award is made to the University of Wisconsin-Milwaukee (UWM) to support the development of multimodal instrument for simultaneous mechanical and fluorescence spectroscopy measurements of single molecules and molecular aggregates. This instrument will enable cutting-edge research and education in the areas of molecular and cellular biology, high-throughput drug screening and cellular analysis, and development of new biomaterials. This new instrument will be part of the Biophysical Microspectroscopy Facility and will serve several laboratories at UWM, as well as other institutions locally, nationally and abroad. Building on their strong record of involving underrepresented minorities in STEM and training graduate, undergraduate and high school students in their research programs, the PIs will use this project to also educate the next generation of scientists and engineers. During the development phase, undergraduate, graduate and postdoctoral researchers will be involved in the design, custom-manufacturing, computer programming, and development of new protocols for imaging, single molecule spectroscopy and for data analysis. This project will also involve middle school and high school students from the Milwaukee area, and will be used for demonstration purposes and for pilot data acquisition and analysis, respectively. Once completed, this facility will provide novel capabilities to the research programs of numerous early-career and senior investigators; it will also play an essential role in preparing the technologically literate workforce of the future, through elaborate outreach programs, as well as teaching and training of undergraduate, graduate, and post-graduate students that will run during the award period and beyond. The new instrument design will be made available to other researchers or US companies interested in bringing this technology to market.
The instrument developed here combines magnetic tweezers, a technique developed by the PI to trap paramagnetic beads and apply mechanical forces to single molecules and cells with Forster resonance energy transfer (FRET) and fluorescence spectroscopy, a method developed by the co-PI to study interactions between molecules and aggregates with spectral resolution. Such a hybrid instrument is not currently commercially available and will require several labor-intensive steps, such as design and fabrication of custom parts, optical alignment and optimization, an integrated control software interface, etc. This instrument presents some unique features. The parallel implementation of force and fluorescence spectroscopy will allow measurements of single-molecule fluorescence with spectral resolution of about 3 nm (over most of the visible spectrum) under mechanical perturbation with a force resolution of 0.1 pN up in the nN range and over hour-to-days sampling times. The multidisciplinary team assembled by the PI is ideally suited to develop and validate this technology, and combines exquisite technical expertise, facilities, experience with developing and running an imaging facility, and close collaborations between physicists, biologists and other life scientists. The instrument will be used for a variety of research studies that involve lateral interactions between proteins under controlled static or dynamic force situations and freely diffusing molecules of a different or the same type, e.g., to study force-induced ligand binding, folding intermediates, antibody detection at single molecule level, receptor-receptor association, and receptor-ligand interactions. The project is jointly funded by the Office of Integrative Activities Major Research Instrumentation Program and the Molecular Biophysics Cluster in the Division of Molecular and Cellular Biosciences.
This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.
