Microplate assays provide information on whole cell populations rather than tracking the behavior of individual cells. Life Technologies offers microplate assays for whole cells and assays performed on disrupted cells or cell lysates. Metabolic activity is commonly used as a viability indicator, but for some applications it can be important to assess viability independent of metabolic state and appropriate assays are listed below. In some cases, cells are required for additional downstream functional analysis and alamarBlue® is an excellent non-toxic indicator of viability.

Select microplate assays for cell viability

See also cell viability assays for flow cytometry
See also cell viability assays for imaging

Measuring Cellular Reduction Potential

  • Simple and scalable, from single well to HTS
  • Accurate and sensitive down to 50 cells

The inherent reducing power of live cells serves as an indicator of metabolic activity and thus cell viability; when cellular reduction potential is measured in a population, the signal is proportional to the number of live cells. Some established assays such as MTT use that reducing power to generate a strongly colored reporter, formazan. Similarly, the redox indicator resazurin is converted to the fluorescent and colorimetric reporter molecule resorufin in metabolically active cells and it serves as a very reliable and sensitive viability/cytotoxicity indicator.

Life Technologies makes the assays simple and convenient by providing the reagents in a proprietary stabilizing formulation, with a mix, incubate, and read protocol that is scalable from single wells to high-throughput screening (HTS).

 alamarBlue® Reduction
Monitoring the efficacy of a growth inhibitor with alamarBlue® indicator.

Measuring Membrane Permeability

  • Measure viability independent of metabolic activity
  • Multi-parameter assays for live and dead cells

The loss of plasma membrane integrity is a key marker of cell death that is regularly exploited by tools that measure viability and cytotoxicity. Cells with compromised plasma membranes allow the influx of cell-impermeant DNA-binding dyes that fluoresce only when bound to DNA. Thus, DNA staining can be used as a cytotoxicity indicator that is independent of metabolic state. Cytotoxic compounds can damage cells through a range of mechanisms. Cell viability, membrane integrity, and DNA content are among the most specific and sensitive parameters for measuring cytotoxicity and provide excellent sensitivity and accuracy.

A richer result is generated when DNA binding is combined with measurements of other parameters. Probes for esterase activity or cellular reduction potential stain live cells and provide measures of viability. A cell-permeant DNA stain will also stain cells with intact plasma membranes and serve as a marker for viable cells.

 HCS LIVE/DEAD® Green Kit

Dose response for valinomycin in HeLa cells using the HCS LIVE/DEAD® Green Kit.

Cell viability can be determined using microplate assays to measure indicators metabolic activity.

 
TargetMeasures cellular reduction potential where signal is proportional to the number of live cells
ReadoutFluorescence or absorbance readoutColorimetric readout in cell lysates
Fluorescent labelResazurin reduced to resorufinXTT/FormazanMTT/Formazan
Ex/Em (nm)571/585 ABS ABS
BibliographyCitationsCitationsCitations
MultiplexingNoNoNo
PlatformMicroplateMicroplateMicroplate
Format25 mL100 mL100 mg1 kit/10 plates
Cat. No.DA1025DAL1100X6493V13154

Cell viability can be determined using microplate assays to measure indicators metabolic activity or esterase activity. Cell membranes increase their permeability in damaged and dying cells allowing the influx of dyes which can both be measured using fluorescence changes. Combination assays measure multiple parameters to provide more context for determining the status of both live and dead cells.

 
TargetDetects membrane permeability using fluorescent DNA binding dye(s)
Single-color assayTwo-color assayAlso measures metabolic activityAlso measures esterase activity
ReadoutDead cells fluoresce greenIntact cells emit mostly green, and dead cells emit mostly redDead cells emit mostly green, and active cells emit mostly redActive cells emit mostly green, and dead cells emit mostly red
Fluorescent labelImage-iT® DEAD Green™Image-iT® DEAD Green™C12-resazurinCalcein-AM
NuclearMask™ Deep Red or Hoechst 33342SYTOX® GreenEthidium homodimer-1
Ex/Em (nm)488/515488/515563/587488/535
638/686 or 350/461504/523528/617
BibliographyCitationsCitations
MultiplexingYesYesNoNo
PlatformMicroplateMicroplateMicroplateMicroplate
Format1 vial/25 plates1 kit/2 plates1 kit/10 plates1 kit/10 plates
Cat. No.I10291H10290L34951L3224