This equipment application is to upgrade our Flow Cytometry capability at the Cleveland VA Medical center. Currently 3 main laboratories (Anthony, Canaday, Shive) have benefitted over the past 6 years from a 2014 BD LSRFortessa 5 laser (Blue, Red, Violet, Yellow/Green, UV) 16 color, Flow Cytometer. We now have an increase in VA funded large volume flow cytometry users (7 main users in present application, including Anthony, Canaday, Shive, Wald, King, Hise, Zidar and with 1-2 additional incoming users anticipated in 2020). The proposed A5 Symphony 5 Laser/ 28 detector (color) configuration flow cytometer will allow the users to move to a higher parameter top end instrument. Symphony advantages include: 1) highest performing commercial fluorochrome based flow cytometer, with similar operation parameters and interfacing software so that current users can transition to easily. 2) A5 comes with 5 lasers, but can be upgraded to 9 lasers as available fluorochromes extend. 3) A5 has higher detector capability, making for increase from current 16 color limit to up to 28 color limit without more lasers 4) Faster processors 5) Better optimized PMT for each detector The end-result of the proposed will accomplish 2 main goals: 1) Allow for cell resource conservation allowing more studies on a given volume of blood by combining flow panels into a single tube. As an example, a single flow panel with 28-colors can provide information on B cells, dendritic cells, and monocytes, optimized for cryopreserved peripheral blood mononuclear cells. Being able to achieve the highest number of studies on limited blood volumes and cell numbers is particularly important in the COVID-19 studies since actual cell yields are reduced in these patients and achievable blood volumes are also diminished due to their critical illness and the longitudinal sampling that we desire. 2) Allow for higher phenotype resolution A number of well described 28 color flow panels (published, and ones proposed within) can provide remarkable depth of analysis for analysis of T cells, iNKT and NK, dendritic cells, and checkpoint pathway focus. The proposed BD A5 Symphony will include: ? 5 Laser/ 28 Color A5 FACS Symphony 5 laser (Sapphire 488-200, OBIS 637-140, Violet 405-200, OBIS 561-150, UV 355-60) Flow Cytometer (allowing 28 colors) ? High Throughput System (HTS) ? HP Z2G4 Win10 w/ 34? monitor ? Extra optical filters for BV510 and V450 ? Fluorochrome & Prototype Dye evaluation kit ? HP Panel Design Starter Kit (50 small vials which included off the shelf, custom, or Optibuild) ? FACS Flow Supply System ? Quantiflash LED Pulsar
Multi-parameter flow cytometric analysis is a key methodology of most studies involving cellular immunology, in both mouse and human systems. The characterization of complex cellular processes, immune regulation, and fine cell subset definition has evolved to include requirement for detecting the presence or absence of an increasing array of cell surface and intracellular markers. The proposed BD Symphony A5, while currently planned to house 5 lasers, will have an option of adding up to 4 additional lasers as needed/indicated as technology advances. It will increase our flow cytometry capability from 16 color flow currently to 28 color flow cytometry. For these reasons we have identified the BD A5 Symphony as the ideal instrument to meet our flow cytometry needs over the coming 5 years, embracing high throughput advanced technology in a way our users can best benefit. Published algorithms and current user expertise will make this transition a natural one.
