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Viability/Cytotoxicity Assay Kit for Animal Live & Dead Cells

The Viability/Cytotoxicity Assay Kit for Animal Live & Dead Cells provides green/red fluorescent staining of live and dead cells, respectively. The assay employs two probes that detect intracellular esterase activity in live cells and compromised plasma membrane integrity in dead cells.

30002 Calcein AM EthDIII-icc cmyk
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30002, - 30002-TView allHide

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150 assays, 300 assays

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Product Description

The Viability/Cytotoxicity Assay Kit for Animal Live & Dead Cells provides green/red fluorescent staining of live and dead cells, respectively. The assay employs two probes that detect intracellular esterase activity in live cells and compromised plasma membrane integrity in dead cells. The esterase substrate calcein AM stains live cells green, while the membrane-impermeable DNA dye Ethidium homodimer III (EthD-III) stains dead cells red. The kit is suitable for detection using fluorescence microscopy, fluorescence microplate reader, or flow cytometry. The assay principles are general and applicable to most eukaryotic cell types, including adherent cells and certain tissues, but not to bacteria or yeast. This fluorescence-based method of assessing cell viability can be used in place of trypan blue exclusion, 51Cr release, and similar methods for determining cell viability and cytotoxicity. EthD-III (#40050 and #40051) is a proprietary DNA stain developed by Biotium to be a superior alternative to the spectrally similar dye Ethidium homodimer I (EthD-I). EthD-III has higher DNA binding affinity and thus brighter fluorescence (30% brighter than EthD-I).

Spectral Properties
Calcein (end product after hydrolysis of calcein AM):
Ex/Em: 494/517 nm (pH 8)

Ethidium homodimer III (EthDIII):
Ex/Em: 530/620 nm (with DNA)

References

1. Biotechniques 29, 874-880 (2000).

 

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MTT and XTT are colorimetric based assays, while resazurin can be measured using colorimetric or fluorescence detection. MTT is not a soluble product, so the cells must be lysed to solubilize the formazan salt before absorbance can be measured. XTT and resazurin do not require cell lysis, allowing kinetic monitoring of the same samples at different timepoints.

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Check to make sure your cell densities are in the linear range of the cell viability assay. Too few cells may fall below the limit of detection, and too many cells may saturate the OD. Perform an initial experiment to check the best linear range for your cell type and experimental set-up. You may need to vary the cell density assay incubation time. The kit protocols provide general guidelines and may need to be optimized for your experimental system.

Category: Apoptosis, Necrosis, and Cell Viability Kits

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Resazurin is reduced by cells to the fluorescent product resorufin. Resorufin can be reduced further to a non-fluorescent compound. Therefore the densest wells may have lowest fluorescence due to over-reduction of the substrate. Please see the product information sheet for more details. The kit protocol provides general guidelines and may need to be optimized empirically for your experimental system. You may need to vary cell density or assay incubation time to ensure that your samples fall in the linear range of the kit.

Category: Apoptosis, Necrosis, and Cell Viability Kits

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Shipment Method: Shipping and handling methods will be assessed and calculated at time of shipment based upon item(s) storage temperature conditions. Expedited shipment may be requested at time of checkout. Please note that products with recommended storage at 4°C or -20°C may ship at ambient temperature. This will not affect product performance. When you receive the product, place it under the recommended storage conditions.