The specific aim of this proposal is to modernize the cell sorting and multi-laser flow cytometry analysis/sorting instrumentation available to the investigators. Secondarily, the presence of this new instrumentation will benefit the entire research community at the University of Illinois at Urban-Champaign (UIUC). Currently, the investigators utilize a 13 year old, heavily modified, EPICS 751 cell sorter operated by the UIUC Biotechnology Center flow cytometry core facility. The EPICS 751 was originally acquired through a NIH SIG in 1985. It has served as a reasonably reliable, heavily used, cell sorting workhorse. Significant effort has been made to extend the life of this instrument by adding items such as: Cicero sorting electronics; improved optics; and additional lasers (all funded by UIUC). The EPICS 751 no longer meets the investigators' research demands for reliable cell sorting and multi- laser flow cytometry analysis. The EPICS 751 is also no longer eligible for service contract support from the manufacturer. The investigators propose that the NIH build on the success of the previous SIG and partially fund the purchase of a MoFlo model MLS2000 cell sorter to replace the aging EPICS 751. The proposed instrument will provide an excellent shred instrument platform, adaptable to the diverse needs of the major user group. Examples of specific uses by the investigators include: selection of rare yeast mutants genetically engineered to express T cell receptor or single-chain antibody fragments (Kranz); selection of transfected primary cells or cell lines based on ectopic plasmid expression to study regulation and pathways of adhesion (Horwitz); multi-color sorting of myeloma cells based on CD45 expression to study the interaction among PI 3-kinase, insulin receptor substrate-1 and CD45 in the inducement of apoptosis (Freund); sorting of hematopoietic progenitor cells into 96 well plates for single cell PCR analysis to the study of the role of the IGF-1 receptor in apoptosis (Kelly_; multi-laser sorting of memory and naive T ells to study effects of aging on the immune system (Woods).
