Table 1.

Comparison of available pharmacological assays for measuring cell viability

Cell viability assayType of assayProsConsSuitable for HTSUsed ina
CellTiter-Glo (Promega)Viability, membrane integrity, ATPNontoxic, does not require fixing or washing stepsATP could be reduced by metabolic interferences, leading to false-positive results; underestimation of drug effect by DNA-targeting agents, in which cells may be incapacitated but intactYesCCLE, GSK, FIMM, SU2C, NTP
Syto60 (Invitrogen)Proliferation; fluorescent DNA stainSimpleRequires fixation, washing; could stain DNA of intact but not-viable cellsYesCGP
Bromodeoxyuridine-ELISA (CytoSelect, Cell BioLabs)Proliferation, DNA synthesisOnly live and replicating cells can incorporate bromodeoxyuridineAssay is time consuming and cost prohibitive in high-throughput formatNo
DAPIProliferation, nucleic acids stainSimpleCould stain nonviable cells if DNA is intactYesGSK
MTT, MTS, XTTViability, metabolic reduction of tetrazolium dyeSimple colorimetric assayFormazan product insoluble in aqueous media; secondary step required to solubilize it before optical detectionNoNCI60
SRBProliferation, anionic general biomass stainSimple colorimetric assay; stable for extended periods; differentiates cell kill from growth inhibitionRequires fixation, washing stepsYesNCI60
Resazurin (CellTiter-Bue; Promega)Viability, cell permeable redox indicatorSimple and inexpensive; more sensitive than tetrazolium assaysPotentially cytotoxic; requires long incubation 1–4 h; could underestimate toxic effectYes
GF-AFC (CellTiter-Fluor; Promega)Viability; penetrates live cells where cytoplasmic cytopeptidase activity releases AFC that fluoresces; protease becomes inactive upon cell deathNontoxic, does not require fixing or washing steps; short incubation time <1 hPotential underestimation of drug effect by DNA-targeting agents, in which cells may be incapaciated but intactYes
Ruthenium dyeProliferation; the fluorescence of the dye is quenced by oxygen; proliferating cells reduce external oxygen and increase fluorescenceNontoxic, simple, rapid assayNarrow dynamic range; signal depends on metabolic activityYes
LDH (Cytotox-ONE; Promega)Cytotoxity; released LDH is measured by enzymatically coupled reagent chemistryReagents compatible with viable cells; quick incubation; cost effectiveSusceptible to background signal from serum sources of LDH in growth media or from compounds inhibiting LDH activityYes
  • aCCLE (3); CGP (4); FIMM, Institute for Molecular Medicine, Finland (38); GSK, GlaxoSmithKline Study (17); NCI60, National Cancer Institute (39); NTP: National Toxicology Program (40); SU2C, the Lawrence Berkeley National Laboratory (41).