Cell viability assay


Cell viability is a measurement of how healthy a cell is. A few ways one can measure cell viability includes looking at apoptosis, cell cycle, or even metabolism, depending on the research goals. There are various methods i.e. flow cytometry, immunocytochemistry, microscopy, etc.



It is critical to understand the degree of cell death in any flow cytometry assay and exclude those cells from the analysis. For live/dead cell discrimination in flow cytometry, BioLegend provides DNA dyes, Helix NP™ NIR, DRAQ7™Propidium Iodide and 7-AAD, that enter and stain dead cells, but are impermeable to live cells for rapid, cost-effective analysis of unfixed cells. In cases where cell fixation is required, BioLegend provides fixable Zombie Aqua™, Zombie Green™, Zombie NIR™, Zombie Red™, Zombie Violet™, Zombie UV™, and Zombie Yellow™.



Zombie dyes exploit the permeability of the cell membrane to indicate live versus dead, but do not bind to DNA. Zombie dyes are a family of amine-reactive dyes that do not passively cross the cell membrane due to their valency. Only if the cell is compromised can these small molecules passively cross. Inside the cell, they covalently conjugate an abundance of intracellular proteins and the signal is retained with paraformaldehyde fixation. Titration of this process is critical since the goal is to use the lowest possible concentration of the Zombie dye to achieve a bright intracellular staining (dead cell status) with low residual cell surface staining (live cell status/background staining). Zombie dyes are most commonly used in flow cytometry, but when optimized, can also be applied to imaging cell in culture.



In experiments where observation of cell health and proliferation by microscopy is required, BioLegend provides Zombie Green™Zombie Red™, and Zombie Violet™.



Not sure which Zombie dye is right for you? Try our Zombie Fixable Viability™ Sampler Kit, which contains 100 tests each of the UV, NIR, Violet, Aqua, and Yellow variants.



Tracking cell division


Proliferation of cells can be measured using probes that are retained by daughter cells, nuclear markers that are only expressed by cycling cells, or assays that assess metabolic activity. BioLegend provides a portfolio of kits and reagents for tracking cell proliferation.

  • CFSE - Retained in daughter cells following cell division, can be used for ex vivo and in vivo cell proliferation assays. Its peak excitation and emission wavelengths are 492 nm and 517 nm, respectively.
  • Tag-It™ Violet - Retained in daughter cells following cell division, can be used for ex vivo and in vivo cell proliferation assays. Its peak excitation and emission wavelengths are 395 nm and 455 nm, respectively.
  • BrdU - Incorporated into newly synthesized DNA, and can be detected with anti-BrdU antibodies.
  • Phase-Flow™ BrdU Kits - Complete set of reagents for analyzing BrdU incorporation by flow cytometry. Kit includes BrdU pulsing solution, anti-BrdU antibody, all necessary buffers, and DNA dyes DAPI and 7-AAD.
  • Ki-67 - Nuclear protein expressed only in cycling cells, and not found in quiescent or senescent (G0) cells.
  • Deep Blue Cell Viability™ - Measures resazurin reduction to determine cell metabolic activity and rate.
  • LDH-Cytox Assay™ Kit - Measures tetrazolium salt reduction resulting from lactate dehydrogenase (LDH) activity. Since LDH is released from damaged cells, this assay can be used to determine cytotoxicity.


To study cell proliferation using flow cytometry, CFSE is widely used for proliferation assays and in vivo cell division tracking. CFSE is able to passively diffuse into cells. Inside the cell, its acetate groups are cleaved by intracellular esterases, and the molecules are converted to fluorescent esters. CFSE is retained within the cell and covalently couples to intracellular molecules via its succinimidyl group. Due to this covalent coupling reaction, fluorescent CFSE can be retained within the cell for an extremely long period. Also, due to this stable linkage, once the dye has been incorporated within the cell, it is not transferred to adjacent cells. The CFSE Cell Division Tracker Kit is composed of lyophilized CFSE and anhydrous DMSO.

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