When you sit down to design your flow cytometry experiment, Cell viability is often relegated to a secondary element. Unfortunately, this decision can drastically affect to your results.
- Don’t let non-viable cells mess up your experiment
- Fix your cells forever with eFluor® Fixable Viability Dyes and live happily
- It’s better to use Fixable Viability Dyes than 7-AAD Viability Dye and Propidium Iodide (PI)
Don’t let non-viable cells mess up your experiment
Excluding dead cells from data is recommended for all staining protocols to ensure accurate data. Dead cells can be “sticky”, resulting in non-specific binding, which causes high background staining and false positives. Moreover, gating based on forward and side scatter (FSC/SSC) is not a reliable method for distinguishing dead cells since Perfetto et al described it back in 2006… At this point, Fixable viability dyes show up to help you, but how?
Fixable Viability dyes ensure dead cells are removed from analysis, thereby reducing non-specific binding and background in addition to better peak separation.
See that cloud in the upper side? Take all the inviable off and the population changes
eBioscience offers several options for ensuring only live cells are analyzed:
- DNA-intercalating dyes:7-AAD, Propidium Iodide (PI)
- Enzymatically sensitive dyes
- Fixable viability dyes: eFluor® Fixable Viability Dyes
Fix your cells forever with eFluor® Fixable Viability Dyes and live happily
eBioscience eFluor® Fixable Viability Dyes (FVD) penetrate compromised membranes, irreversibly labeling dead cells from all species prior to fixation and permeabilization. Dead cell populations have high fluorescent intensity, allowing for easy exclusion, thereby improving data quality. Fixable Viability Dyes eFluor® 455UV, eFluor® 450, eFluor® 506, eFluor® 520, eFluor® 660 and eFluor® 780 are permanent dyes suitable for UV, violet, blue and red lasers that will not wash out of cells. Unlike 7-AAD and propidium iodide, cells labeled with Fixable Viability Dyes can be cryopreserved or fixed, permeabilized and stained for intracellular antigens without losing staining intensity of the dead cells.
Reasons we love Fixable Viability Dyes:
- Ready to use format
- Easy to exclude unviable cells
- Data improvement
It’s better to use Fixable Viability Dyes than 7-AAD Viability Dye and Propidium Iodide (PI)
As mentioned before, 7-AAD Viability Dye and Propidium Iodide (PI) Staining Solution mark non-viable cells by intercalating with the DNA of dead cells in a concentration-dependent manner. The nucleic acid of viable cells will not be accessible to the dye, thereby preventing staining. They are not recommended for viability gating if the cells will be fixed. 7-AAD and PI may also be used for flow cytometric analysis of the cell cycle. PI binds to the double stranded DNA of dead cells, but is excluded from cells with intact plasma membranes.
Labclinics has a special offer: 25% OFF in ALL FIXABLE VIABILITY DYES Invitrogen Ebioscience, valid until March, 31st 2017.
For further information, contact us at tecnic@labclinics.com and we’ll be happy to help.
References
Perfetto SP, Chattopadhyay PK, Lamoreaux L, Nguyen R, Ambrozak D, Koup RA, Roederer M. Amine reactive dyes: an effective tool to discriminate live and dead cells in polychromatic flow cytometry. J Immunol Methods. 2006 Jun 30;313(1-2):199-208. Epub 2006 May 19.
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