This is a proposal to acquire a MoFlo Astrios flow cytometer/high-speed cell sorter to replace an old MoFlo Legacy (2001), increasing capacity and adding to capabilities of the current cell sorting facility, a key component for 10 major NIH-funded research programs. The rationale for this request rests on a careful assessment based on the following factors: 1. A need to improve cell sorting service capacity in the Flow Cytometry Facility as crucial for the current NIH-funded investigators and the anticipated growth in Wistar Institute programs. 2. One of 2 sorters, the FACSAria II, is capable of 16 fluorochrome/4 laser detection while the existing MoFlo allows for only five fluorochromes. Upgrading would allow greater flexibility and efficiency for investigators moving from multi-color analysis to sorting, not restricting them to only one sorter. 3. Experiments requiring bio-safety containment at levels greater than BSL-1 are currently restricted to the FACSAria II, resulting in scheduling bottlenecks. The proposed new cell sorter would be appropriately equipped to allow for BSL-2+ sorting. 4. Fluorescent protein sorting is currently limited, given the lack of appropriate laser excitation. The proposed sorter would have a laser in the green/yellow range for optimal excitation of fluorescent proteins such as mCherry, mTomato, etc. 5. The proposed sorter would be capable of six-way sorting, providing the ability to separate more populations than the current four, along with the ability to sort into slides or plates up to 1536 wells. 6. Te existing MoFlo Legacy is still capable of performing well and used on a mostly daily basis, and is preferred by many for its speed and sample recovery, but is suffering from more frequent and increasing amounts of downtime due to age-related repairs. This results in increasing amounts of operator time and places NIH-funded projects at risk due to the potential for longer downtimes due to availability of only one sorter. While major overhaul and enhancement is an option, the expense involved would be better applied into a new instrument. This well-regarded facility, established over 30 years ago, is currently staffed by three experienced technicians and monitored by a faculty user committee. Many investigators at Wistar and neighboring institutions routinely use the cell sorting services, with the existing sorters often heavily booked with occasional backlogs due to capacity limitations as discussed above. Therefore, we propose to upgrade sorting capacity to maintain state- of-the-art support for 10 major and 8 minor users, with 41 R01/P01/U01 NIH-funded research awards, through purchase of a well-equipped MoFlo Astrios high-speed sorter, with appropriate bio-safety level containment, six-way sorting, and the ability to sort single cells directly into tissue culture plates or slides.
|Allegrezza, Michael J; Rutkowski, Melanie R; Stephen, Tom L et al. (2016) IL15 Agonists Overcome the Immunosuppressive Effects of MEK Inhibitors. Cancer Res 76:2561-72|
|Facompre, Nicole D; Harmeyer, Kayla M; Sole, Xavier et al. (2016) JARID1B Enables Transit between Distinct States of the Stem-like Cell Population in Oral Cancers. Cancer Res 76:5538-49|
|Zhang, Ying; Ertl, Hildegund C J (2016) Depletion of FAP+ cells reduces immunosuppressive cells and improves metabolism and functions CD8+T cells within tumors. Oncotarget 7:23282-99|
|Allegrezza, Michael J; Rutkowski, Melanie R; Stephen, Tom L et al. (2016) Trametinib Drives T-cell-Dependent Control of KRAS-Mutated Tumors by Inhibiting Pathological Myelopoiesis. Cancer Res 76:6253-6265|
|Shannan, Batool; Watters, Andrea; Chen, Quan et al. (2016) PIM kinases as therapeutic targets against advanced melanoma. Oncotarget 7:54897-54912|