DESCRIPTION (provided by applicant): Funds are requested for the purchase of a Tecnai G2 Spirit BioTWIN G2 transmission electron microscope (TEM) and microtome. This TEM is a state-of-the-art instrument and contains a high-contrast BioTWIN objective lens optimized for 2D and 3D imaging of stained or unstained, low-contrast samples. All of the microscope components including the electron gun, the optical elements, the vacuum system and the stage, are digitally controlled, allowing the user to automatically recall all optimized operating conditions including lens settings, gun parameters, optical alignments, aperture alignments. Similarly, the CCD camera is computer controlled so that image acquisition can be automated and connected to a network for exporting data. Four major users and seven minor users, whose work is supported by 16 RO1's, and 3 R21's, 1 PO1 and 1 DP2, will utilize the microscope. These users will account for approximately 32 hr/week (80% usage of the microscope time). The remainder of the time will be available for new users at Schepens Eye Research Institute (SERI) and, when available, for outside users. The requested TEM will replace the current a Phillips 41OLS, which is 25 years old and for which replacement parts are no longer available. The Tecnai G2 Spirit BioTWIN G2 lists at $659,492 but we have negotiated a very significant discount and have a quote to purchase the scope, camera and tomography stage at $600,000. The high internal usage of the scope makes it infeasible to use services at neighboring institutions. The scope will be integrated into the Morphology Unit and will be overseen and operated by Ms. Patricia Pearson, who has been an integral part of our Morphology Unit for over 30 years and is recognized as an expert in the field of ocular histology. A fulltime research technician, supported by SERI, who will be responsible for tissue preparation and sectioning, will assist Ms. Pearson. Work on the TEM will be carried out as a fee-for-service, with work done on a first come-first serve basis. The service contract will be with Nanotech, who specializes in the service of Philips and FEI electron microscopes, and who has been servicing our current scope for more than 20 years. The cost of the contract is built into the fees that are charged for the TEM services. SERI also maintains a fund for the service and maintenance of common equipment and these funds will be available for expenses not covered in any contract, as was done with the Phillips 41OLS. An internal advisory committee will assist the Principal Investigator with maximizing the utilization and overseeing the day-to-day management of the microscope as well as with establishing a financial plan for the long-term operation and maintenance of the microscope. Dr. Dennis Brown who heads the Imaging Core for the Mass General Program in Membrane Biology has been added to the advisory committee and will be available for advice on novel applications such as tomography. Institutional commitment is evidenced in the included letter from Dr. Eli Peli, a Co- Director of Research at SERI. As more and more of our research has come to rely on ultrastructural visualization of cells and tissues, demand for the TEM has grown and access to this state of the art TEM will undoubtedly facilitate and support our work. ) PUBLIC HEALTH RELEVANCE: Transmission electron microscopy (TEM) permits the visualization of cells and tissues at the subcellular level by providing very high magnification. There has been an increasing utilization of imaging technologies to understand the structure/function of genes of interest. The requested TEM will be used to examine how certain genes and proteins are involved in normal visual processes and how they are altered in diseased tissues of the eye.

Transmission electron microscopy (TEM) permits the visualization of cells and tissues at the subcellular level by providing very high magnification. There has been an increasing utilization of imaging technologies to understand the structure/function of genes of interest. The requested TEM will be used to examine how certain genes and proteins are involved in normal visual processes and how they are altered in diseased tissues of the eye.