Calculate blank-corrected relative cell viability or method-specific percent alamarBlue reagent reduction. Choose the result your control design supports: an untreated-cell control and a fully reduced reagent control serve different purposes.

Enter sample, cell-free background and untreated-cell fluorescence. Optional sample replicates replace the single sample reading with their mean.

Keep relative viability against untreated cells separate from reagent reduction against a fully reduced positive control. Readings must share reagent concentration, incubation and instrument settings. Out-of-range results are reported, not clamped; check backgrounds, saturation and assay interference.

Optional settings

Comma, space or newline separated; matched background/control settings are required.

Choose the appropriate method

Relative viability = 100 × (sample RFU − blank RFU) / (untreated-control RFU − blank RFU). This expresses signal relative to untreated cells; it is not the percentage of reagent reduced. Optional sample replicates replace the sample reading with their mean.

Fluorescence reduction = 100 × (sample RFU − blank RFU) / (100%-reduced-control RFU − blank RFU).

For paired absorbance, reduction = 100 × (Eox,2A₁ − Eox,1A₂) / (Ered,1C₂ − Ered,2C₁). A values are test absorbances; C values are cell-free medium-plus-reagent negative controls. Do not pre-subtract C from A.

Manufacturer coefficients
Wavelengths (nm)Eox₁Eox₂Ered₁Ered₂
570 / 6008058611721615567714652
570 / 63080586347981556775494
540 / 6004761911721610439514652

Worked example

Sample 600 RFU, blank 100 RFU and untreated control 1,100 RFU give 100 × 500 / 1,000 = 50% relative viability. If the fully reduced reagent control is instead 2,100 RFU, the reduction result is 100 × 500 / 2,000 = 25%.

Frequently asked questions

Why can results exceed 100%?

A stronger signal than the reference gives a value above 100%. The calculator keeps it visible; check background, interference and assay range.

What does the replicate SD describe?

Sample variability only. The control and blank are treated as fixed, so it is not a confidence interval or total assay uncertainty.

Can fluorescence be converted to absorbance?

Not with a universal factor. Use the measurement protocol and its required controls.

Reference: Thermo Fisher alamarBlue user guide, fluorescence and paired-absorbance calculations.