Flow Cytometry Core Facility
flow cytometry

Instrumentation

The UK Flow Cytometry and Cell Sorting Core Facility has two analytical flow cytometers: a Becton-Dickinson LSRII and a Becton-Dickinson FACSCalibur. The facility also has two iCyt high-speed cell sorters, which are housed in a biosafety cabinet within a separate biocontainment room. The facility also has an automated magnetic cell sorter, the Miltenyi Biotec AutoMACS system.

LSR II

This analyzer can simultaneously measure up to 8 fluorescent colors, as well as forward light scatter (cell size) and side light scatter (cell granularity). It is equipped with 3 lasers: 488nm and 633nm lasers and a 355nm UV laser.

LSR II

FACSCalibur

This cell analyzer can simultaneously measure up to 4 fluorescent colors, as well as forward light scatter (cell size) and side light scatter (cell granularity). It is equipped with two lasers: 488nm and 633nm.

FACSCalibur

iCyt Synergy sorter system

This flow cytometer system contains two separate sorter units housed in a biosafety laminar flow hood. Each sorter unit is equipped with 5 lasers (355 nm, 405nm, 488nm, 561nm and 642nm) and can sort or analyze cells labeled with up to 12 antibodies or fluorescent labels. In addition, the biocontainment setup allows sorting and analysis of infectious samples (risk groups 1 and 2) and live human samples. Cells can be sorted sterilely into either tubes or microtiter plates.

iCyt

AutoMACS Magnetic Sorter

Sorting on this instrument utilizes antibodies that are labeled with an iron-containing compound, which are then separated automatically from unlabeled cells with a rare earth magnet. Cells are labeled either directly or indirectly with conjugated antibody and then are placed on the magnetic sorter for automatic separation of labeled cells. Cells can also be labeled at the same time with a fluorochome-labeled antibody and subsequently run on a flow cytometer to check purity. The AutoMACS system from Miltenyi-Biotech is a powerful enrichment tool when combined with the flow cytometers when trying to purify a rare cell type. Labeled antibodies for use in the AutoMACS sorter can be purchased through the Miltenyi website:  http://www.miltenyibiotec.com/en/default.aspx.

AutomMACS

Data Analysis

Standardized data analysis is available to investigators for each set of samples, depending upon the application. Data can be reanalyzed at the request of the investigator with a 30-minute minimum service charge. The flow facility also makes available several workstations and software options for users to analyze their own data at no additional charge. The workstations in the lab have either Mac-based CellQuest analytical software or PC-based Summit software as well as FlowJo software. Alternatively, free analytical software can be downloaded to the investigators office/lab, including WinMDI for PCs and FCS Assistant for Macs. FlowJo software for either Mac or PC computers is available for users to purchase at the following website: http://www.flowjo.com/index.php.