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| Product Name | Propidium Iodide (PI) |
| Catalog No. | RESK-0527 |
| Description | Propidium iodide (PI) is a widely used red fluorescent nuclear stain. As an analog of ethidium bromide (EB), it intercalates between bases to bind DNA with little or no sequence preference, binding approximately one dye molecule per 4-5 DNA base pairs. PI also binds RNA, so nuclease (RNase) treatment is required to distinguish DNA from RNA staining. In aqueous solution, PI has excitation/emission maxima of 493/636 nm. Upon binding to nucleic acids, the fluorescence signal is enhanced 20-30 fold; the excitation maximum shifts ~30-40 nm toward the red and the emission maximum shifts ~15 nm toward the blue, giving excitation/emission maxima of 535/617 nm. Although PI has a relatively low molar absorptivity, its large Stokes shift allows simultaneous detection of nucleic acids and fluorophore-labeled antibodies when appropriate filters are used. PI is suitable for fluorescence microscopy, confocal microscopy, flow cytometry, and fluorometry. PI is membrane-impermeant and excluded from live cells, but it passes through compromised membranes to stain the nucleus. Based on this property, it is commonly combined with live-cell fluorescent probes such as Calcein-AM, Hoechst 33258, or Hoechst 33342 to simultaneously stain and identify live and dead cells in apoptosis-related research. PI can also serve as a counterstain in multiplex fluorescence staining and is compatible with a variety of cell-labeling techniques, including direct and indirect fluorescent antibody detection, mRNA in situ hybridization, cell-structure-specific fluorescent probes, and tissue staining. PI staining alone can also be used for cell cycle analysis. |
| Intended Use | Nuclear staining of dead or membrane-compromised cells; live/dead cell discrimination in apoptosis research (with live-cell probes); counterstaining in multiplex fluorescence labeling; cell cycle analysis by flow cytometry. For research use only. |
| Principle / Technology | PI intercalates between DNA bases (about one dye per 4-5 base pairs, with little or no sequence preference). Because PI cannot cross intact cell membranes, it selectively stains dead or membrane-damaged cells. Nucleic-acid binding enhances fluorescence 20-30 fold and shifts Ex/Em from 493/636 nm to 535/617 nm. |
| Detection Method | Fluorescence microscopy Confocal microscopy Flow cytometry (FL2 channel) Fluorometry |
| Sample Type | Cultured cells (live/dead discrimination), fixed cells, and tissue samples |
| Performance Range / Specifications | English synonym: 2,7-Diamino-9-phenyl-10-(diethylaminopropyl)-phenanthridium iodide methiodide CAS NO.: 25535-16-4 Formula: C27H34I2N4 Molecular weight: 668.4 Appearance: dark red crystals Purity: ≥95% by HPLC Solubility: soluble in water (1 mg/mL) Fluorescence spectra: in aqueous solution, PI Ex/Em = 493/636 nm; PI-nucleic acid complex Ex/Em = 535/617 nm; fluorescence can be excited by a xenon lamp, a pump arc lamp, or a 488 nm argon-ion laser; typically detected in the FL2 channel of a flow cytometer |
| Specificity | Stains nucleic acids with little or no sequence preference (one dye per 4-5 DNA base pairs); also binds RNA. Excluded from live cells with intact membranes; stains nuclei of dead or membrane-compromised cells. |
| Reaction Conditions / Protocol | Dissolve in water (soluble at 1 mg/mL) to prepare a stock solution and dilute to the working concentration with an appropriate buffer. For DNA-specific staining, treat samples with RNase to remove RNA. |
| Components / Formulation | Supplied as a solid (dark red crystals). |
| Storage Conditions | Store at 4°C, protected from light. |
| Shelf Life | Stable for at least one year when stored at 4°C protected from light. |
| Package Specifications | 10 mg 100 mg |
| Product Form | Dark red crystals |
| Quality Control | Purity ≥95% by HPLC. |
| Purity | ≥95% by HPLC |
| Molecular Weight | 668.4 |
| Shipping Conditions | Shipped on wet ice. |
| Expiration Date / Stability | Stable for at least one year at 4°C protected from light. |
| Regulatory / Compliance | For research use only. |
| Compatibility | Compatible with live-cell probes such as Calcein-AM, Hoechst 33258, and Hoechst 33342 for simultaneous live/dead cell staining. Compatible with direct and indirect fluorescent antibody detection, mRNA in situ hybridization, cell-structure-specific fluorescent probes, and tissue staining. Excitable by xenon lamp, pump arc lamp, or 488 nm argon-ion laser; detected in the FL2 channel of flow cytometers. |
| Recommended Buffer System | Aqueous buffers; soluble in water at 1 mg/mL. |
| Application Notes / Precautions | PI is membrane-impermeant: it is excluded from live cells and stains only cells with compromised membranes; combine with live-cell probes for live/dead discrimination in apoptosis research. PI also binds RNA; use RNase treatment when DNA-specific staining is required. FAQ - Why does RNase treatment follow formaldehyde/paraformaldehyde/glutaraldehyde fixation but is usually unnecessary after methanol/acetic acid or acetone fixation? Cross-linking fixatives (formaldehyde, paraformaldehyde, glutaraldehyde) inactivate enzymes during cross-linking, so RNase must be added afterwards to remove RNA. Methanol/acetic acid or acetone fixation works by denaturation with only mild protein effects and allows re-solubilization after fixation, so RNase treatment is generally not required. Protect from light. |
For research use only, not for clinical use.
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