NucRed® Dead 647 ReadyProbes® Reagent Protocol for Fixed Cells
Nuclear stain for fixed cells
NucRed® Dead 647 ReadyProbes® Reagent emits bright far-red fluorescence when bound to DNA and is suitable for staining nuclei in fixed-cell preparations and tissue sections. NucRed® Dead 647 reagent may also be used to measure viability and to measure cytotoxicity in kinetic live-cell assays.
This protocol can be used for:
- Nuclear staining to identify nuclear condensation in apoptotic cells using fluorescence microscopy
This protocol should not be used for:
- Flow cytometry
You will need the following for this protocol:
- Cells growing in culture
- NucRed® Dead 647 ReadyProbes® Reagent (Cat. No. R37113)
- Fluorescence microscope with far red or Cy®5 filters
Labeling fixed cells
First, fix and permeabilize cultured cells with a protocol appropriate for your sample.
|1. Wash the cells 1–3 times in PBS as needed.|
|2. Add two drops of NucRed® Dead 647 Reagent per milliliter of medium.|
|3. Incubate for 5–15 minutes, protected from light.|
|4. Remove the stain solution.|
|5. Wash the cells 2–3 times in PBS.|
|6. Image the cells.|
|Standard filter set||Cy®5|
|EVOS® Light Cube||Cy®5|
|Storage conditions||Room temperature|
- In some cases, more or fewer drops may be needed to achieve optimal staining intensity.
- Image quality may be improved by replacing the culture medium with Live Cell Imaging Solution (Cat. No. A14291DJ).
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.
For Research Use Only. Not for use in diagnostic procedures.