Fixable Viability Dye eFluor® 660 is a viability dye that can be used to irreversibly label dead cells prior to cryopreservation, fixation and/or permeabilization procedures. Unlike 7-AAD and propidium iodide, cells labeled with Fixable Viability Dyes can be washed, fixed, permabilized, and stained for intracellular antigens without any loss of staining intensity of the dead cells. Thus, using Fixable Viability Dyes allows dead cells to be excluded from analysis when intracellular targets are being studied. These dyes may be used to label cells from all species. Fixable Viability Dye eFluor® 660 can be excited by the red (633 nm) laser line and detected using standard filter sets for APC or Alexa Fluor® 647 (660/20 band pass filter). Please make sure that your instrument is capable of detecting this dye. For compensation, it is recommended to use a sample of the cells of interest stained with the Fixable Viability Dye. If the percentage of dead cells is expected to be less than 5%, then it is recommended to take a small aliquot of cells and heat them at 65°C for 1 minute then immediately place on ice for 1 minute. After this treatment, the heat-killed cells can be combined 1:1 with live cells and then stained with the Fixable Viability Dye. Fixable Viability Dye eFluor® 660 is supplied as a stock solution prepared in high-quality, anhydrous DMSO. It should be protected from light and moisture. Store at -80°C with dessicant. It may be freeze-thawed up to 20 times. Allow the vial to equilibrate to room temperature before opening.