This proposal requests the purchase of a LUMICKS C-Trap instrument for a Major User group at the University of Vermont (an IDeA state institution). The C-Trap is the only commercially available laser trap with combined scanning confocal fluorescence microscopy. Mechanobiology is an emerging field concerning physical forces and their impact on biological functions. In cells, tiny molecular motors interact with tracks to generate force and motion involved in biological movement critical to muscle contraction, intracellular cargo transport (such as insulin granules), and DNA repair. Therefore, investigators utilize biophysical approaches to correlate the structural dynamics (nm motions in ms timescales) and mechanics (pN forces) of individual molecular motors and protein machines as they undergo critical protein-protein interactions with their tracks while performing their normal physiological functions. The LUMICKS C-Trap would provide University of Vermont investigators the capability to make such measurements at the single molecule level. Although a custom-built laser trap exists at the university, it's 21 years old with simple epifluorescence; preventing investigators from performing true molecular structure and functional studies described above. More importantly, the existing laser trap is built with components no longer supported or manufactured; placing users at risk of a total system failure. The requested LUMICKS C-Trap with its real-time correlative, laser trapping and scanning confocal fluorescence microscopy, is a turn-key system with a multi-channel microfluidic experimental chamber. This chamber makes setup of single molecule protein-protein interaction assays relatively simple. Couple this ease with the C-Trap's user friendly software control and data collection/analysis package, investigators productivity should be greatly enhanced. In addition, the mechanistic insights governing the processes that the molecular machines studied undergo will become apparent given the unprecedented resolution of molecular structure and function made possible with the LUMICKS C-Trap. Four Major Users have been identified for the purpose of this application as they represent most of the projected demand (i.e. 80%) for the requested instrument, given their existing and proposed experimental requirements. We also identified a strong Minor User group that will use the remaining time on the proposed system. These two University of Vermont User groups represent faculty and biomedical research interests from the Departments of Molecular Physiology & Biophysics, Pharmacology, and Microbiology & Molecular Genetics. These faculty also represent the Cardiovascular Research Institute, Vermont Cancer Center, and the Cellular, Molecular, and Biomedical Sciences and Biomedical Engineering Graduate Programs. The LUMICKS C-Trap with its management through the College of Medicine's Microscopy Imaging Center will afford the capacity for advanced single molecule mechanical recording and structural imaging and ensure sustained and efficient use for the university's biomedical research community for the foreseeable future.
The goal of this proposal is to purchase a LUMICKS C-Trap microscope that will allow scientists to probe the most basic molecular processes that cells in the body use to create movement and force, which underly muscle contraction and insulin secretion, as examples. Therefore, mutations to the proteins, associated with these normal and critical processes, can lead to human diseases such as genetic forms of heart failure, cancer, and diabetes.
