Membrane integrity–based viability assay

NucRed Dead 647 ReadyProbes Reagent is a cell-impermeant stain that emits bright far-red fluorescence when bound to DNA. Since this reagent stains cells that lack plasma membrane integrity, it is extremely useful to measure viability and to measure cytotoxicity in kinetic live-cell assays. NucRed Dead 647 reagent is also suitable for staining nuclei in fixed-cell preparations and tissue sections.

This protocol can be used for:

  • Identifying dead cells using a fluorescence microscope

This protocol should not be used for:

  • Flow cytometry

You will need the following for this protocol:



1. Culture cells in appropriate medium and vessel for microscopy


2. Add 2 drops NucRed® Dead 647 reagent per milliliter of medium to label cells


3. Incubate 5–15 minutes


4. Image cells

Spectral information and storage

 NucRed Dead 647 ReadyProbes Reagent
Excitation/Emission642/661 nm
Standard filter setCy5
EVOS Light CubeCy5
Storage conditionsRoom temperature

 Protocol tips


Tubulin and nuclear staining in HeLa cells. HeLa cells were fixed and permeabilized and stained for tubulin (biotinylated mouse anti-a-tubulin mouse monoclonal antibody followed by an Alexa Fluor 488 goat anti-mouse secondary antibody and counter-stained with NucRed Dead 647 ReadyProbes reagent.