Hexidium iodide, a fluorescent nucleic binding acid stain (excitation/emission ~ 518/600 nm), permeants to mammalian cells and selectively stains almost all gram-positive bacteria. Hexidium iodide can bind to the DNA of all bacteria after permeabilization by EDTA.
In Vitro
Hexidium iodide allows assessment of Gram status by differential absorption through bacterial cell walls, selectively staining gram-positive organisms in suspension. MCE has not independently confirmed the accuracy of these methods. They are for reference only.
Form:Solid
Especificaciones
Especificaciones y pureza
≥98%
Mecanismos bioquímicos y fisiológicos
Hexidium iodide, a fluorescent nucleic binding acid stain (excitation/emission ~ 518/600 nm), permeants to mammalian cells and selectively stains almost all gram-positive bacteria. Hexidium iodide can bind to the DNA of all bacteria after permeabilization
Condiciones de almacenamiento de almacenamiento
Store at 2-8°C,Protected from light,Desiccated
Enviado en
Wet ice
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Incubate with 1–5 µg/mL dye for 10–20 min; wash and mount in antifade medium.
Notes
Work under subdued light. Prepare fresh working solutions. Concentrations and times are starting points; titrate for best S/N on your microscope or cytometer.
Biological Roles
Hexidium iodide is a synthetic fluorescent probe and does not have a natural biological role. Its interactions with biological systems are primarily physicochemical (literature/general):
Nucleic-acid binding: Intercalates between DNA/RNA base pairs, stabilizing the duplex locally and enhancing red fluorescence upon binding.
Membrane interactions: As a cationic, relatively bulky molecule, its cellular uptake is governed by membrane permeability; Gram-negative bacterial outer membranes typically restrict entry, enabling Gram-type discrimination (literature).
Photophysics in cells: Fluorescence intensity and spectral profile depend on local environment (viscosity, polarity, nucleic-acid density). Photobleaching can occur under intense illumination.
Cytotoxicity: Like many intercalating dyes, prolonged exposure or high concentrations can be cytotoxic; assays are therefore designed to minimize exposure and use controls (literature). No clinical or therapeutic use is implied.
Conclusion: Its utility arises from selective accumulation and fluorescence enhancement upon nucleic-acid binding, making it a valuable research probe for imaging and cytometry.
Buffer Applications
This product is not a buffering reagent. However, buffer selection is critical for successful staining and detection (literature/general guidance):
Recommended media: Use neutral, isotonic buffers such as PBS or HEPES-buffered saline without phenol red to minimize background fluorescence.
Protein/serum effects: Serum proteins and polyanions (e.g., heparin) can bind cationic dyes and alter staining; consider protein-free buffers for consistent results.
Divalent cations: Mg2+/Ca2+ can influence membrane permeability and dye uptake in bacteria; maintain consistent ionic conditions across samples.
pH: Maintain pH 7.2–7.6 for live-cell staining; extreme pH can quench fluorescence or degrade the dye (literature).
Sterility: For flow cytometry or live-cell imaging, sterile-filter working solutions (0.22 µm) to avoid contamination.
Note: No item-specific buffer composition is provided for this SKU; adjust buffer systems empirically to your assay and verify compatibility with your instrument’s optics.
Green Alternatives
While hexidium iodide is effective and convenient, some laboratories consider alternative dyes with improved environmental, health, or waste profiles.
Potential alternatives (literature/general):
SYBR Safe or GelGreen (nucleic acid stains): Designed for reduced mutagenic potential compared with ethidium-like dyes; suitable for gels and some staining workflows. May differ in permeability/selectivity vs bacterial membranes.
EvaGreen/Brilliant dyes: High-sensitivity DNA-binding dyes for qPCR and solution assays with lower reported hazard profiles; not direct functional substitutes for Gram differentiation.
Propidium iodide (PI): Widely used red nucleic-acid stain; however, hazard profile is similar to other intercalating dyes and not necessarily greener.
Comparison summary (literature; qualitative):
Hexidium iodide: Strong red emission; Gram-positive selectivity; excited near 488 nm.
SYBR Safe/GelGreen: Safer handling claims by manufacturers; typically green emission; different excitation/emission and permeability properties.
EvaGreen: Low-background, high-sensitivity in solution/qPCR; limited use for Gram-selective imaging.
Trade-offs:
Greener dyes may require different instrument channels, provide different selectivity, or need re-optimization of staining protocols.
Waste considerations:
Collect dye-containing waste separately; consult institutional guidance on disposal of intercalating dye waste irrespective of dye chosen.
Pharmaceutical Uses
Not applicable as a pharmaceutical or excipient. Hexidium iodide is supplied strictly for research use only and is not intended for human or veterinary use, diagnostics, or clinical applications.
Fluorescent probes like hexidium iodide are occasionally used in research-stage process analytics (e.g., cell-based assay development, microbiological monitoring) but are not components of finished dosage forms.
No pharmacopeial monograph is known for this dye class; there are no established excipient specifications for hexidium iodide.
Always follow the Research Use Only notice and do not incorporate into products intended for administration.
Physical Properties
Item-specific specifications (this lot):
Melting/boiling point, density, refractive index, solubility, logP, pKa: Not specified for this item; refer to CoA/Spec Sheet.
Fluorescence and related properties (literature/general; matrix-dependent):
Absorption (unbound): broad in blue–green; practical excitation with 488 nm lasers has been reported (literature).
Emission (bound to nucleic acids): red/orange fluorescence typically detected ~600–650 nm; exact maxima depend on solvent, binding state, and environment (literature).
Fluorescence enhancement: substantially increases upon intercalation into DNA/RNA relative to aqueous solution (literature).
Solubility tendencies (literature/general):
Readily soluble in polar media such as water and DMSO; aqueous stocks are commonly prepared for immediate use, while DMSO stocks can be more stable and easier to aliquot (literature). Actual solubility limits depend on grade and counterion content.
Photostability and light sensitivity (literature/general):
Light-sensitive fluorescent dye; prolonged exposure to light can reduce signal due to photobleaching. Work under subdued light.
Note: Use the product-specific CoA/SDS for authoritative physical and safety properties. Literature values above are general guidance for the class of dye and may vary with conditions and formulation.
Quality and Grades
Grade/Purity: Not specified for this item; refer to CoA/Spec Sheet.
Context for fluorescent dye quality (general guidance):
Dye content and counterion balance: The effective concentration depends on chromophore purity and stoichiometric iodide content. Reputable QC includes HPLC/UPLC purity, water content, and identity by MS/UV-Vis.
Spectral QC: UV-Vis absorption profile and fluorometric response upon DNA binding are often used to verify performance. Low background fluorescence in aqueous media is desirable.
Impurities: Trace colored/fluorescent impurities can increase background and reduce signal-to-noise. Metal contaminants are typically not a primary concern for use as a stain, but residual solvents/moisture can affect solubility.
Batch-to-batch consistency: Particularly important for quantitative flow cytometry or high-content imaging workflows. Use the CoA to confirm spectral characteristics and any recommended working concentrations for the supplied lot.
Stabilizers and formulation:
Stabilizer/preservative content: Not specified for this item; refer to CoA/Spec Sheet.
Takeaway: Rely on the item’s CoA for exact purity metrics and any recommended performance criteria. Validate dye performance in your assay matrix upon receipt.
Reaction and Applications
This product is primarily a fluorescent probe rather than a synthetic reagent. Accordingly, its most relevant applications are analytical/biological assays (research use only).
Analytical and imaging applications (literature/general):
Bacterial differentiation: Enters and strongly stains Gram-positive bacteria; Gram-negative cells typically exclude it due to the outer membrane, enabling differential staining in mixed populations.
Viability/permeability assays: When paired with membrane-permeant green dyes (e.g., SYTO family), enables two-color discrimination by microscopy or flow cytometry.
Nucleic acid staining: Intercalates into DNA/RNA with enhanced red fluorescence upon binding; used in fixed-cell imaging and nucleic-acid gel/slide staining workflows.
Flow cytometry: Excitable at 488 nm; detected in PE-Texas Red or PerCP-Cy5.5–like channels depending on instrument filters (literature; verify on your cytometer).
Practical tips (literature/general):
Work under subdued light. Prepare fresh working dilutions just before use to minimize hydrolysis/photobleaching.
Optimize concentration empirically; too high a concentration can raise background or quench fluorescence.
Include appropriate controls: unstained, single-stained compensation controls, and dye-minus-one controls for multicolor panels.
Note: No manufacturer application notes were provided for this SKU; the above reflects common research practices with hexidium iodide–type dyes.
Reaction Conditions
Conventional reaction-condition guidance (solvent, catalysts, temperature) is not applicable because this product is used as a fluorescent probe, not as a reagent.
Stock solutions: Commonly prepared in DMSO (e.g., 1–10 mM) or water at high µg/mL to mg/mL levels; filter-sterilize if used with live cells. Actual solubility and stability depend on lot and solvent.
Working concentrations: Typical final ranges span ~1–10 µg/mL for microscopy or flow cytometry; titrate for your cell type and instrument sensitivity.
Incubation: 5–20 minutes at room temperature is often sufficient for bacterial staining; minimize light exposure. For fixed samples, longer incubations may be used.
Detection: Excite with 488 nm where available; collect emission with 600–650 nm bandpass filters appropriate for your optics (literature; verify settings on your system).
Controls: Include unstained, single-color controls, and viability controls (e.g., heat-killed or permeabilized cells) to interpret permeability-dependent staining.
These parameters are general literature practices and should be optimized empirically for each assay and instrument.
Safety and Handling
Authoritative data for this item:
GHS classification, hazard statements (H-phrases), signal word, and pictograms: Not specified for this item; refer to SDS.
General laboratory safety guidance for cationic intercalating dyes (literature/good practice):
Hazards: Many nucleic acid intercalators can be harmful if swallowed, inhaled, or absorbed through skin and may cause eye/skin irritation. Avoid generating dust or aerosols.
PPE: Use lab coat, nitrile gloves, and safety glasses or face shield. Change gloves frequently when handling concentrated dye stocks.
Handling: Work under low light to minimize photobleaching; avoid contact with skin and surfaces because of intense staining. Use secondary containment for stock solutions.
Incompatibilities: Strong oxidizers and strong reducing agents may degrade the dye; avoid high pH/bleach solutions when preparing stocks. Do not autoclave dye solutions (risk of degradation and pressure build-up in sealed vials).
First aid (overview; defer to SDS): In case of skin contact, wash with plenty of water and soap. For eye contact, rinse cautiously with water for several minutes and seek medical attention. If inhaled, move to fresh air. If ingested, rinse mouth and seek medical attention.
Spill/cleanup: Absorb small spills with inert material (e.g., cellulose pads), collect for disposal. Decontaminate with appropriate detergent; avoid bleach with concentrated dye as it may produce fumes.
Always consult the product SDS for definitive hazard classifications and emergency measures.
Solvent Selection
Hexidium iodide is a polar, cationic dye typically handled as concentrated stock solutions.
Polarity/miscibility (literature/general):
Water: Generally soluble; useful for immediate-use working solutions. Ionic strength and pH can influence spectral properties and binding.
DMSO: Highly soluble; convenient for preparing stable, concentrated stocks and for aliquoting to minimize freeze–thaw.
Alcohols (MeOH/EtOH): Moderate solubility; may be used for cleaning or intermediate solutions.
Nonpolar solvents: Poorly soluble; not recommended.
Practical selection guidance:
Use DMSO stocks (e.g., 1–10 mM, literature) to enable accurate micro-volume dispensing; dilute into aqueous buffers immediately before use to final micromolar or microgram-per-milliliter levels.
Choose protein- and phenol red–free buffers (e.g., PBS, HEPES) to reduce background. Serum proteins can bind cationic dyes and alter staining.
Avoid strong bases and oxidants that degrade the chromophore.
Comparison (literature trends):
DMSO vs water stocks: DMSO offers improved shelf stability and easier aliquoting; water stocks are simpler but may have shorter stability and higher risk of microbial growth (unless sterile-filtered).
Note: No item-specific solvent specifications are provided; consult the CoA for any restrictions applicable to this lot.
Storage and Reconstitution
Item-specific storage from Product Data:
Store at 2–8°C.
Protect from light.
Keep desiccated.
Shipped on wet ice.
Reconstitution and handling (general literature guidance; not item-specific specifications):
Solvents: Prepare concentrated stocks in anhydrous DMSO or water. DMSO stocks are often preferred for long-term integrity and accurate microliter dispensing.
Concentration: Common stock levels are 1–10 mM (DMSO) or high µg/mL–mg/mL (aqueous). Filter-sterilize (0.22 µm) if using with live cells.
Aliquoting: Divide into single-use aliquots to minimize light exposure and repeated freeze–thaw.
Working solutions: Prepare immediately before use in appropriate buffer (e.g., PBS or HEPES) and discard unused portions at end of day.
Stability notes:
Maintain the product container tightly closed with desiccant; minimize headspace moisture.
Avoid repeated warming/cooling cycles. Keep protected from ambient light at all times.
For any extended storage of reconstituted solutions, consult the SDS/CoA for this item. Literature practice for similar dyes is to store aliquots at ≤–20°C, protected from light; however, follow the item’s labeled storage (2–8°C) for the supplied material unless your internal validation dictates otherwise.
Always refer to the product’s CoA and SDS for definitive storage limits, shelf life, and compatibility.
Structure and Identity
Brief overview: Hexidium iodide is a cationic, red-fluorescent, nucleic-acid–binding dye of the ethidium/phenanthridinium family, supplied as the iodide salt. It is widely used for bacterial differentiation and membrane-permeability assessments in research settings.
SKU: H650905
Product name: Hexidium iodide
CAS: 211566-66-4
Category: Small molecules and compound libraries (research use only)
InChIKey: Not specified for this item; refer to CoA/Spec Sheet.
SMILES: Not specified for this item; refer to CoA/Spec Sheet.
Molecular formula: Not specified for this item; refer to CoA/Spec Sheet.
Molecular weight: Not specified for this item; refer to CoA/Spec Sheet.
Structural features (general/literature):
Quaternary, permanently charged aromatic heterocycle (phenanthridinium/ethidium-like chromophore) paired with iodide as counterion (literature).
Extended, polycyclic aromatic system that intercalates between base pairs of DNA/RNA; positive charge promotes binding to polyanionic nucleic acids (literature).
Typically lacks stereocenters; planar cationic chromophore with auxochromes that red-shift emission upon nucleic-acid binding (literature).
2D description in words (literature):
A fused tricyclic aromatic cation (phenanthridinium-type) bearing dialkylamino substituents; counterbalanced by iodide. The planar aromatic cation enables intercalation; the quaternary nitrogen confers permanent positive charge.
Synthetic Utility
This material is a finished probe, not a building block or reagent for bond-forming synthesis. Therefore, traditional synthetic utility (e.g., as a catalyst, electrophile, or protecting group) is not applicable.
Relevant chemistry notes (literature/general):
Functional groups: Quaternary ammonium (phenanthridinium-like) cation; polycyclic aromatic system; iodide counterion. These features underlie its photophysical properties and nucleic-acid intercalation rather than synthetic reactivity.
Stability considerations: The aromatic cation can undergo photobleaching or oxidative degradation; harsh nucleophiles/bases may attack auxochromic substituents.
If chemical modification is desired (e.g., to make conjugates), practitioners typically start from appropriately functionalized phenanthridinium/ethidium scaffolds rather than from hexidium iodide itself.
Target Specificity
Antibody/biomolecule target specificity does not apply. However, as a fluorescent probe, hexidium iodide exhibits the following binding preferences (literature/general):
Primary target: Nucleic acids (DNA and RNA) via intercalation into stacked base pairs.
Selectivity: Enhanced uptake/staining in Gram-positive bacteria compared with Gram-negative due to outer membrane exclusion in the latter. Within nucleic acids, many phenanthridinium dyes show modest preference for double-stranded regions and GC-rich sites, though sequence specificity is not absolute.
Cellular localization: Predominantly nucleoid in bacteria and nuclei/nucleoli in eukaryotic cells once membranes are permeabilized or in fixed preparations.
Note: Binding and permeability are strongly assay- and organism-dependent; confirm selectivity with appropriate biological controls.
Preguntas frecuentes
What is the purity of this product?
This product is supplied at ≥98% purity (chemical assay). Lot-specific values are stated on the Certificate of Analysis.
How should this product be stored?
Store at 2–8 °C, protected from light, desiccated. Keep the container tightly closed with desiccant — the material is moisture-sensitive.
How is this product shipped?
This product ships chilled on wet ice. Unpack on arrival and transfer it to the storage condition stated above.
What are the CAS number, molecular formula and molecular weight?
The CAS Number is 211566-66-4, the molecular formula is C25H28IN3, and the molecular weight is 497.41 g/mol.
What documentation is provided?
Available product documentation, including Certificates of Analysis (COA), Safety Data Sheets (SDS), and specification sheets, is shown in the product document area. Document availability and access follow the current site policy.
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