Flow Cytometry cell cycle analysis
direct, convenient analysis of cell cycle...
On single laser machines, DRAQ5™ can be combined with R-PE & FITC conjugated antibodies, in a simple protocol with no compensation .....
At stoichiometric concentrations DRAQ5™ can provide cell cycle information (eg %G0/1, %S, %G2/M) in mixed populations identified by FITC / PE labelling.
Incubation of antibody stained samples takes only ten minutes with DRAQ5™.
Using an appropriate software (e.g. Cylchred, available from Cytonet UK) it is possible to generate cell subset specific cell cycle plots as shown below. Recent reports confirm that data using DRAQ5™ approaches the data quality of propidium iodide and ethidium bromide. Take a look at review of papers confirming this by Plander et al (2003), Yuan et al (2004), Primo (2006) and Swerts (2007).
EXAMPLE:
Burkitt's Lymphoma sample stained with anti-CD3-FITC (FL1), anti-CD20-PE (FL2) and DRAQ5 (FL3). FL1 is plotted against FL2. FL3 is plotted (Area vs Width) to exclude doublets (R3). Typically the R1/R3 analysis (T cells) will show quiescent cells i.e. G0/G1 while the R2/R3 analysis will have a significant number of B lymphoma cells in S and G2/M phases.Data courtesy of Stephen Couzens, Haematology, University Hospital of Wales, Cardiff.
DRAQ5™ can additionally provide S-Phase Fraction (SPF) data, one measure of proliferation, which can be useful in some diseases e.g. breast cancer. Again, this can be achieved on cell subsets.
DRAQ5™ has also been utilised on the laser scanning cytometer to generate cell cycle data on drug treatment and hypoxia (Schwock et al, 2005 ). Use the left navigation and go to Applications>Laser Scanning Cytometry to view this.
Research applications of DRAQ5™-based cell cycle analysis are numerous. See Weng et al for an example with live GFP transfectants and controls.
DRAQ5™ can be used with FITC / PE stained samples for the rapid labelling of nucleated cells in a 'no lyse/no wash' protocols. This can provide a rapid screening test for use in situations where scatter information is not required for interpretation.
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