A group of nine NIH-funded major users and four secondary NIH-funded users at Fox Chase Cancer Center request funds for a BD Biosciences LSR flow cytometer. The currently available instrumentation that is capable of providing >3 color analysis, a BD FACStar' plus cell sorter is at capacity, particularly due to increasing demands for sorting. The other instrument, a BD FACScan, is widely used for simple immunophenotyping (2 color, P1) and for cell cycle analysis, but cannot perform more complex multi-color analyses that are increasingly required by use of various fluorescent reporter systems (e.g., IRES-GFP retroviruses). Furthermore, many of the users in this application propose experiments that currently cannot be done with the present instrumentation, specifically determination of intracellular calcium levels or DNA content together with multicolor cell surface markers. The proposed instrument is a three-laser analyzer that will be user-operated, providing for maximum flexibility in scheduling and providing increased multicolor analysis capacity together with these new capabilities. While the research programs of the different users are quite diverse, including immunobiology, virology, and cell biology, they all have as an ultimate goal the understanding of human disease processes. This is being accomplished by studies of the mechanisms of lymphocyte development, imrnunoglobulin rearrangement, generation of functional subpopulations of effector cells (such as memory B cells, helper T cells, cytotoxic T cells, and NK cells), regulation of the cell cycle, signal transduction, and regulation of cell morphology. Some of the proposed experiments analyze viruses that infect the central nervous system. Others seek to increase our understanding how immune responses, both humoral and cellular, can be regulated, with implications for vaccination and also autoimmunity. Still others have a more clinical focus, seeking to develop a vaccine therapy for certain cancers. All of these projects have a common need for making multiple precise measurements of individual cells, typically in complex mixtures, a requirement that can be met effectively by the proposed instrument. The requested instrument will be housed and maintained in the Flow Cytometry Facility at FCCC. Users will be trained in its operation by Facility personnel. This shared Facility is an ideal location for this system as it provides Flow Cytometry technical expertise and instrument access to NIH funded laboratories at FCCC and is a hub of collaborative efforts with outstanding, highly trained technical support.