SYTO® 82 Nuclear Staining Protocol
Nuclear stain for eukaryotic and prokaryotic cells
SYTO® 82 is a cell-permeant, non-exclusive nucleic acid stain that shows a large fluorescence enhancement upon binding. In both live and dead eukaryotic cells, SYTO® 82 generally shows cytoplasmic or mitochondrial as well as nuclear staining. In addition, SYTO® 82 will stain most live and permeabilized bacteria.
This protocol can be used for:
- Nucleic acid (nuclear) staining in fluorescence microscopy
This protocol should not be used for:
- Flow cytometry
You will need the following for this protocol:
- Cells growing in culture
- SYTO® 82 Orange Fluorescent Nucleic Acid Stain (Cat. No. S11363)
- Fluorescence microscope
|1. Culture cells in an appropriate medium and vessel for fluorescence microscopy.|
|2. Remove the medium.|
|3. Wash the cells 1–3 times in a phosphate-free buffer to remove the medium.|
|4. Prepare the SYTO® 82 staining solution by diluting the stock solution 1:1,000 (5 µM) in a phosphate-free buffer.|
|5. Add sufficient staining solution to cover the cells.|
|6. Incubate for 5–30 minutes, protected from light.|
|7. Remove the staining solution.|
|8. Wash sample 3 times in a phosphate-free buffer.|
|9. Image the cells.|
|Standard filter set||TRITC|
|EVOS® Light Cube||RFP|
- Warm to room temperature and briefly centrifuge the DMSO solution to the bottom of the vial each time before use.
- Try multiple dye concentrations in the range from 100 nM to 5 µM to determine the optimal concentration.
- In general, the best results are obtained in buffers that do not contain phosphate, such as Hank’s Balanced Salt Solution (Cat. No. 14025092).
Cells stained with SYTO® 82 dye and imaged with the Thermo Scientific™ CellInsight™ High-Content System.
For Research Use Only. Not for use in diagnostic procedures.