Researchers at the University of Pennsylvania are increasingly engaged in investigations that require greater than six-parameter flow cytometry. In addition, many investigators are making use of fluorescent resonance energy transfer (FRET) experiments to map receptor interactions. Currently, these types of multiparameter studies must be performed on the cell sorters (only 1 capable of measuring FRET with the preferred CFP/YFP pair) as no analytical instrument capable of performing sophisticated multiparameter analyses is available. Since these instruments are also used for preparative cell sorting, the wait for analytical only applications is often weeks. Lack of access to high-speed 6+ parameter analytical flow cytometry has proven to be a serious limitation to several well-funded investigators. The Becton- Dickinson LSR II (BD LSRII) is an air-cooled four-laser bench top cytometer capable of measuring more than 12 parameters simultaneously. Using a new digital acquisition system, it has enhanced detection and resolution capabilities. As configured with UV and violet lasers, it will support both calcium flux (with Indo-1) and FRET experiments, which are now available on a very limited basis. These expanded analytical capabilities to the flow cytometry core laboratory will specifically benefit the user group and many researchers in the institution and be a significant addition to the available instrumentation in the School of Medicine. In this proposal, we present a model for shared resource operation that will support the efficient use of the BD LSR II for predominantly the user group. Already in place are significant institutional support, a highly experienced staff, and a well-respected training program in flow cytometry. This will allow investigators immediate access to the service and further extend access by training selected students and technical staff to use. This will result in increased availability of high-level analytical cytometry to the scientific community. By placing the LSR II in existing recently renovated space and utilizing a well-established model for core flow cytometry facility operation, we propose to maximize the benefit of this cytometer to the user group of 17 NIH-funded investigators ($9,007,559 direct costs/year). This will allow investigators to pursue many areas of research, previously with limited availability, which will result in increased funding and advancement of biomedical science.
