This proposal is for fund to a FACStarplus flow cytometer/cell sorter to replace the Ortho 50H now operated by the NYU Flow Cytometry Facility. The Ortho 50H, while remarkably advanced for its time, is an awkward instrument to use as a sorter. In addition to this awkwardness, it is limited in both the number of paramaeters that it can analyze and by the speed at which its 10 year old computers operate. It is also becoming an """"""""orphan"""""""" instrument, poorly supported by its manufacturer. The NYU Flow Cytometry Facility is a core laboratory and provides service to the entire medical center. It is designed to provide """"""""state of the art"""""""" facilities to support more than $7 million in peer review funded research. The FACStarplus requested in this proposal provides the capacity to perform 3 color analysis with the argon laser and 4 color analysis with the combination of argon and dye lasers. Three fluorescence signals can be analyzed (and sorted) with a single laser. One of the lasers has UV capability so that Hoechst dyes can be used for DNA determinations on viable cells and sorting can be performed on the basis of position in the cell cycle. Chromosomal sorting and flow karyotyping are also within the capabilities of this instrument Intracellular Calcium, pH and membrane viscosity determination can also be performed using UV excitation. With the dye pump removed, this laser can be used for fluorescein and phycoerythrin while the other argon laser is used for UV. The 15usec sort cycle permits sorting with high recoveries at rates close to 5000 cells/sec. This is three to four times faster the Ortho. In practical terms this means that a sort designed to produce 5 x 10(7) cells for biochemical analysis or cDNA library production from a source in which the desired cells make up 20% of the population can be performed in 12 hours not two days.
