Covalent labeling of dead cells for flow or microscopy
Live-or-Dye™ Fixable Viability Stains are cell membrane impermeable amine-reactive dyes. The dyes enter dead cells that have compromised membrane integrity and covalently label free amines on intracellular proteins. On live cells, the dyes react with surface proteins, but these are much less abundant than intracellular proteins, resulting in low staining levels compared to dead cells.
Live-or-Dye™ labeling is extremely stable, allowing the cells to be fixed and permeabilized without loss of fluorescence or dye transfer between cells. The staining protocol has been optimized to maximize live/dead discrimination with minimal live cell staining, in order to prevent interference with immunostaining. Biotium offers a selection of ten bright and photostable Live-or-Dye™ stains for maximal flexibility in multi-color analysis. We also offer Live-or-Dye NucFix™ Red, a unique fixable nuclear-specific dead cell stain.
Live-or-Dye™ Fixable Viability Staining Kits are designed for discrimination between live and dead cells during flow cytometry. Live/dead stains are useful probes to include when analyzing cell surface protein expression by flow cytometry, because they allow intracellular fluorescence signal from dead cells with permeable plasma membranes to be excluded from analysis.
The Live-Or-Dye™ Fixable Viability Staining Kits were designed to give a great separation between live and dead cell signals, in order to make it easy to exclude dead cells from your analysis (Fig. 1).
STABLE, FIXABLE STAINING
Fixability is an important feature of a dead cell stain. The ability of a stain to withstand both fixation and permeabilization allows the dye to be combined with intracellular flow cytometry staining protocols or immunofluorescent microscopy experiments, among other applications.
Live-or-Dye™ Fixable Viability Staining Kits have superior fixability and stable staining (Fig. 2). Our bright and photostable dyes allow samples to be fixed, permeabilized, and stored (protected from light) without loss of signal.
It is often very useful to identify dead cells for microscopic analysis as well, particularly when performing live-cell imaging experiments. In microscopy, live/dead stains allow unambiguous visual discrimination of dead cells. Live-or-Dye™ Fixable Viability Staining Kits work just as well for microscopy as they do for flow cytometry: they are bright and photo-stable dyes, with negligible signal in live cells and strong signal in dead cells.
Four stains have been validated for fluorescence microscopy: Live-or-Dye™ 488/515, 568/583, 594/614, & 640/662. The other dyes are expected to work as well, as long as the microscope has the appropriate excitation and emission settings.
LIVE-OR-DYE NUCFIX™ RED
Live-or-Dye NucFix™ Red is a unique, cell membrane impermeable dye that specifically stains the nuclei of dead cells.
Unlike other commonly used nuclear stains such as propidium iodide (PI) or DRAQ7™, Live-or-Dye NucFix™ labeling is covalent, so the dye doesn’t transfer between cells after fixation. Live-or-Dye NucFix™ Red can be used for flow cytometry or fluorescence microscopy.
YEAST-SPECIFIC LIVE-OR-DYE™ STAINING KITS
For yeast and fungal cells we have developed a series of staining kits that combine the dead cell-specific Live-or-Dye™ stains with all-cell stains designed with yeast properties in mind.
The Yeast Live-or-Dye™ Live/Dead Staining Kit combines a cell-permeant green dye, Thiazole Orange, which concentrates in the nucleus of live cells, with Live-or-Dye™ 568/583 (Fig. 5). Co-staining with the two dyes allows live/dead discrimination of yeast by fluorescence microscopy or flow cytometry.
The Yeast Viability Staining Kits take advantage of the unique fungal cell wall, by combining a Live-or-Dye™ dead cell stain with a fluorescent conjugate of the cell wall probe Concanavalin A (Con A) (Fig. 6). These kits are available in 3 color combinations for maximum experimental flexibility.
Live-or-Dye™ Fixable Viability Staining Kits
|32002, 32002-T||Live-or-Dye™ 350/448||355 nm||DAPI or Violet||347/448 nm||Flow Cytometry|
|32003, 32003-T||Live-or-Dye™ 405/452||405 nm||Pacific Blue®||408/452 nm||Flow Cytometry|
|32009, 32009-T||Live-or-Dye™ 405/545||405 nm||AmCyan||395/545 nm||Flow Cytometry,
|32004, 32004-T||Live-or-Dye™ 488/515||488 nm||FITC||490/515 nm||Flow Cytometry,
|32012, 32012-T||Live-or-Dye™ 510/550||488 nm||Spectral scan||516/549 nm||Spectral Cytometry|
|32005, 32005-T||Live-or-Dye™ 568/583||488 or 561 nm||PE||562/583 nm||Flow Cytometry,
|32006, 32006-T||Live-or-Dye™ 594/614||488 or 561 nm||PE-Texas Red®||593/614 nm||Flow Cytometry,
|32007, 32007-T||Live-or-Dye™ 640/662||633 or 640 nm||APC||642/662 nm||Flow Cytometry|
|32013, 32013-T||Live-or-Dye™ 665/685||633 or 640 nm||Spectral scan||667/685 nm||Spectral Cytometry|
|32008, 32008-T||Live-or-Dye™ 750/777||633 or 640 nm||APC-Cy®7||755/777 nm||Flow Cytometry,
|32010, 32010-T||Live-or-Dye NucFix™ Red||488 or 532 nm||PE-Texas Red®||520/593 nm||Flow Cytometry,
Yeast-Specific Fixable Viability Staining Kits
|Catalog No.||Kit Name||Live-or-Dye™ Color||Other Dye|
|31064||Yeast Live-or-Dye™ Fixable Live/Dead Staining Kit||Live-or-Dye™ 568/583 (stains dead cells red)||Thiazole Orange (stains all cells green)|
|31063-1||Yeast Viability Staining Kit (Green/Red)||Live-or-Dye™ 594/614 (stains dead cells red)||CF®488A ConA (stains all cell walls green)|
|31063-2||Yeast Viability Staining Kit (Red/Blue)||Live-or-Dye™ 405/452 (stains dead cells blue)||CF®594 ConA (stains all cell walls red)|
|31063-3||Yeast Viability Staining Kit (Red/Far-Red)||Live-or-Dye™ 568/583 (stains dead cells red)||CF®640R ConA (stains all cell walls far-red)|
Draq7 is a trademark of Biostatus, Ltd. Texas Red and Pacific Blue are registered trademarks of Thermo Fisher Scientific. Cy dye is a registered trademark of GE Healthcare.