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Introduction
PrestoBlue™ Cell Viability Reagent is a ready‐to‐use reagent for rapidly evaluating the viability and proliferation of a wide range of cell types. PrestoBlue™ reagent is quickly reduced by metabolically active cells, providing a quantitative measure of viability and cytotoxicity in as little as 10 minutes. Protect reagent from long term exposure to light and avoid contamination, as this can reduce sensitivity.
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Quick Reference Protocol
- PrestoBlue™ Cell Viability Reagent is supplied as a 10X solution. Add PrestoBlue™ Reagent directly to cells in culture medium. See below for example volumes:
Format Volume of cells + media Volume of PrestoBlue™ Reagent Cuvette 900 μL 100 µL 96-well plate 90 µL 10 µL 384-well plate 36 µL 4 µL Note: Correct for background fluorescence by including control wells containing only cell culture media (no cells) on each plate.
- Incubate ≥10 minutes at 37º C. Longer incubation times will increase sensitivity of detection. As this is a live cell assay, readings may be taken at multiple time points to determine optimal performance in your lab.
Format Recommended Incubation Time Bottom-read fluorescence 10 minutes – 2 hours Top-read fluorescence 30 minutes – 2 hours Absorbance 20 minutes – 2 hours Room temperature incubation 10 minutes – 2 hours Low cell number (< 5,000 cells/100 µL) 20 minutes – 24 hours - Read fluorescence or absorbance. Fluorescence is more sensitive than absorbance and is the preferred detection method.
Format Excitation Emission General 540–570 nm 580–610 nm Fluorescence
(Monochrometer)560 nm (10 nm bandwidth) 590 nm (10 nm bandwidth) Fluorescence (Filter) 535 nm (25 nm bandwidth) 615 nm (10 nm bandwidth) Absorbance 570 nm 600 nm (reference wavelength for
normalization) - Calculate and plot the results. Higher fluorescence or absorbance values correlate to greater total metabolic activity.
Format Instructions Fluorescence - a. Optional: Average the fluorescence values of the no-cell control wells and
subtract from the fluorescence value of each experimental well. - b. Plot fluorescence vs. experimental condition (cell number, compound
concentration).
Absorbance - a. Normalize the 570 nm values to the 600 nm values for the experimental wells.
- b. Plot the normalized 570 nm absorbance values vs. experimental condition (cell
number, compound concentration).
- a. Optional: Average the fluorescence values of the no-cell control wells and
MAN0003232 11–Oct–2010
