Product Name
Propidium Iodide
Product Overview
Propidium iodide (PI) is used for fluorescence imaging of cell nuclei.
Description
Propidium iodide (PI) binds to DNA by intercalation between bases with little sequence preference and a stoichiometry of one dye per 4-5 base pairs of DNA. PI also binds to RNA and requires nuclease treatment to distinguish RNA from DNA staining. Once the dye binds to the nucleic acid, its fluorescence is enhanced 20-30 fold, with the fluorescence excitation maximum shifted about 30-40 nm toward red and the fluorescence emission maximum shifted about 15 nm toward blue. PI is suitable for fluorescence microscopy, confocal laser scanning microscopy, flow cytometry, and fluorescence measurements. PI is commonly used to identify dead cells in populations and as a counterstain in multicolor fluorescence techniques.
Characteristic
Fluorescence excitation and emission profiles of propidium iodide bound to dsDNA.
Usage
Nuclear fluorescent probe
Stability
Stable under normal conditions.
Storage
2-8℃, protect from light.
Introduction
Propidium iodide (PI) is a popular red-fluorescent nuclear and chromosome counterstain. Propidium iodide is commonly used to detect dead cells in a population. This dye is provided as a 10 mg solid and a 1 mg/mL aqueous solution.
Handling
Always wear recommended Personal Protective Equipment. No special handing advice is required.
Category
Plant Organelle Fluorescent Probes
Excitation/Emission (nm)
535/617
Molecular Formula
C27H34I2N4
Counterstaining Cells in Suspension for Flow Cytometry
Sample Preparation
1. Collect a cell suspension of 2 × 105 to 1 × 106 cells.
2. Pellet the cells by centrifugation and discard the supernatant.
3. Tap the tube to resuspend the pellet in the residual liquid and add 1 mL of PBS at room temperature.
4. Transfer the full volume of resuspended cells to 4 mL of absolute ethanol at -20℃ by pipetting the cell suspension slowly into the ethanol while vortexing at top speed. Leave the cells in ethanol at -20℃ for 5-15 minutes.
5. Pellet the cells by centrifugation and discard the ethanol.
6. Tap the tube to loosen the pellet and add 5 mL of PBS at room temperature. Allow the cells to rehydrate for 15 minutes.
Counterstaining Protocol
1. Make a 3 μM solution of PI by diluting the 1 mg/mL (1.5 mM) stock solution 1:500 in staining buffer (100 mM Tris, pH 7.4, 150 mM NaCl, 1 mM CaCl2, 0.5 mM MgCl2, 0.1% Nonidet P-40). A 1 mL volume will be required for each cell sample.
2. Centrifuge the cell suspension from step 6, discard the supernatant, tap to loosen the pellet, and add 1 mL of PI diluted in the staining buffer. Incubate for 15 minutes at room temperature and analyze by flow cytometry in the presence of the dye.
Chromosome FISH Counterstaining
Sample Preparation Prepare the specimen according to standard procedures.
Briefly rinse the final preparations in dH2O before counterstaining to remove residual buffer salts from the slide. This final rinse will help reduce nonspecific background staining on the glass. Allow the preparation to air dry.
Counterstaining Protocol
1. Make a 1.5 μM PI staining solution by diluting the 1 mg/mL (1.5 mM) stock solution 1:1000 in PBS. Pipet 300 μL of this staining solution directly onto the specimen. If necessary, RNase A (freshly made) may be added to a final concentration of 10 μg/mL. A plastic coverslip can be used to distribute the dye evenly on the slide.
2. Incubate the specimen in the dark for 30 minutes at room temperature, or 37℃ if RNase is included.
3. Remove the coverslip and rinse briefly with PBS or dH2O to remove unbound dye.
4. Remove excess liquid from the slide by gently blotting around the sample with an absorbent tissue. Place a glass coverslip on the slide, and seal the edges with wax or nail polish. Alternatively, the preparation can be mounted in an antifade reagent according to the manufacturer’s directions.
5. View the sample using a fluorescence microscope with appropriate filters.
Chemical Name
Propidium iodide
Full Name
PI (Propidium iodide)
Appearance
purple/red solution
Potential Health Hazard Effects
May cause eye irritation with susceptible persons.
May cause skin irritation in susceptible persons.
May be harmful by inhalation.
May be harmful if swallowed.
Polymerization
Hazardous polymerization does not occur.
Notice
Use the reagent as soon as possible after unpacking!
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Datasheet 