Funding is requested for a state-of-the-art high speed fluorescence activated cell sorter to be utilized by a group of seven major users on the Iowa State University campus. The technology utilized in flow cytometry has advanced rapidly in the past five years to include both high speed processing of multi-parameter signals for both analysis and sorting, as well as the incorporation of larger flow cell tip orifices to accommodate the sorting of larger particles (up to 400 micron diameter flow cell tips). The justification for requesting matching funds for the purchase of a high speed sorter can be viewed in three sections. First, the existing EPICS 752 instrument within the facility has been in operation for eight years and has become the limiting factor in the progression of the projects outlined here as a result of increased downtime and the slow rate of data processing for both analysis and sorting. Secondly, many of the research projects have common requirements that the ELITE ESP cytometer will be able to provide which will enable these projects to progress forward. They include the large scale production sorting (> 106 cells) of large cell or particle subpopulations that represent only a small portion of the overall distribution (< 5 %). The following cell or particle types are both large, often fragile, and represent a minor portion of the population to be sorted. They include: swine inner cell mass cells (35 microns), Arabidopsis protoplasts (35 microns), soybean cyst nematode eggs (100 microns), and mouse ine trophoblastic giant cells (200-300 microns). The rapid cell sorting capabilites are also required for the large scale production sorting (> 106 cells) of the traditional smaller mammalian cell types such as Iymphocytes for the purpose of assaying cytokine production and hybridoma cells to be analyzed for the loss of or mutation with the gene encoding the heavy chain for antibody. Additionally, the expansion to a multiple laser system will facilitate the move to four color analyses and cell sorting in the case of the swine Iymphocytes. Finally, the operation of the Facility which will house this instrument has a well documented and solidly financed educational component for training faculty, post doctoral research associates, as well as graduate and undergraduate students in both the experimental design, sample preparation, and use of the fluorescence activated cell sorter in research and the Facility will continue to fulfill this componet with a state-of-the-art instrument. The new ELITE ESP will be used extensively for all of these purposes in both research as well as teaching.
