Cy3 NHS ester bromide , CAS No.2892098-34-7

CAS: 2892098-34-7 Cat. No.: C1450922 Formula: C₃₄H₄₀BrN₃O₄ Molecular Weight: 634.60

Storage

Store at -20°C

Shipped In

Ice chest + Ice pads

Overview

Cy3 NHS ester bromide is a CY dye. CY, short for Cyanine, is a compound consisting of two nitrogen atoms connected by an odd number of methyl units. Cyanine compounds have the characteristics of long wavelength, adjustable absorption and emission, high extinction coefficient, good water solubility and relatively simple synthesis. CY dyes are of en used for the labeling of proteins, antibodies and small molecular compounds. For the labeling of protein antibodies , the combination can be completed through a simple mixing reaction. Below, we introduce the labeling method of protein antibody labeling, which has certain reference significance.

Specifications

Storage

Store at -20°C

Shipped In

Ice chest + Ice pads

This product requires cold chain shipping. Ground and other economy services are not available.

Names and Identifiers

• Molecular Weight: 634.60

Application Protocols

The following protocols are general literature/practice guidelines for NHS-ester Cy3 labeling and are not item-specific validations. Optimize for your system.

A. Antibody labeling (1 mg scale)

• Materials: Antibody (1 mg/mL in 0.1 M NaHCO3, pH 8.3), Cy3 NHS ester stock (10 mM in anhydrous DMSO), desalting column (G-25), PBS.
• Procedure:
  1. Exchange antibody into NaHCO3 buffer (amine-free).
  2. Add dye to 1 mg Ab (≈6.7 nmol) at 3–6 eq (20–40 nmol; 2–4 µL of 10 mM stock). Final DMSO ≤5%.
  3. Incubate 30–45 min at RT in dark with gentle mixing.
  4. Quench with 50 mM ethanolamine, 10 min.
  5. Purify by desalting into PBS. Measure A280 and A550 to compute DoL (use appropriate A280 correction factor for Cy3, literature values ~0.08–0.1; verify empirically).

B. Amine-modified oligo labeling

• Dissolve oligo in 0.1 M NaHCO3, pH 8.5 (50–100 µM). Add 10–20 eq dye from DMSO stock. React 1 h at RT, dark. Purify by spin desalting or RP-HPLC. Confirm by MALDI/ESI-MS and UV–Vis.

C. Amine-functionalized surface labeling

• Swell substrate if polymeric. Treat with dye (1–5 mM) in anhydrous DMF with 1–2 eq DIPEA, 2 h at RT, dark. Rinse with DMF, then alcohol, then buffer. Assess coverage by fluorescence.

General notes: Protect from light, minimize water exposure of dye stock, and verify labeling via spectroscopy and, where appropriate, SDS-PAGE in-gel fluorescence.

Biological Roles

This product is a synthetic fluorescent labeling reagent and does not have an intrinsic biological role in cells or organisms. The following describes typical uses and behavior of Cy3 labels in biochemical contexts (literature/general guidance):

• Function in assays: Cy3 serves as a bright, orange-emitting reporter for tracking biomolecules in vitro. It is commonly used in fluorescence microscopy, flow cytometry, FRET assays (as donor to Cy5), and blotting techniques.
• Conjugation targets: Primary amines on proteins (lysines, N-termini), peptides, amine-modified oligonucleotides, and amine-functionalized surfaces/particles.
• Photophysics: Absorption in green (~548–554 nm) and emission in orange (~565–570 nm) allow excitation with 532–561 nm laser lines. Photobleaching rates depend on oxygen level and local environment; antifade systems and oxygen scavengers can prolong signal.
• Environmental sensitivity: Cyanine fluorescence can be influenced by local polarity, aggregation state, and proximity to quenchers (e.g., tryptophan residues, guanidinium). Protein conjugation may shift λmax slightly.
• FRET: Frequently paired as a donor with Cy5 acceptor in ~5–6 nm Förster distance systems; exact R0 depends on spectral overlap and orientation factor (literature).

Research-use only

• As indicated, this product is for research use only. It is not intended for diagnostic or therapeutic use in humans or animals. No physiological or clinical activity is claimed.

Buffer Applications

This product is not a buffering agent. However, buffer choice is critical during amine-labeling with NHS esters. The following are general, literature-based recommendations:

• Suitable buffers (amine-free):
  • Sodium bicarbonate/carbonate, 50–100 mM, pH 8.3–8.6 (promotes amine nucleophilicity while balancing NHS hydrolysis rate).
  • Sodium phosphate, 50–100 mM, pH 7.4–8.0 for more sensitive proteins (slower reaction; less risk of hydrolysis and denaturation).
  • Borate buffer, 50 mM, pH 8.5–9.0 for robust targets (faster labeling; higher hydrolysis risk).
• Additives: 0.1–0.5 M NaCl can reduce nonspecific interactions; include 1–5% v/v DMSO to aid dye dispersion. Avoid metal chelators only if they interfere with downstream assays.
• Avoid buffers containing primary/secondary amines: Tris, glycine, ethanolamine, HEPES derivatives with amines, or ammonium salts; they compete with the target.
• pH control: Calibrate pH at the working temperature; NHS hydrolysis approximately doubles in rate with each ~10°C rise (rule-of-thumb; literature), so keep reactions at room temperature unless otherwise optimized.
• Quenching: After labeling, residual NHS ester can be quenched with ethanolamine (50 mM) or Tris, added only after the desired reaction time has elapsed.

Note: These are general recommendations; optimize buffer composition empirically for your specific biomolecule.

Green Alternatives

Assessment of current practice

• Conventional solvents: Anhydrous DMSO or DMF are typically required to dissolve Cy3 NHS esters; both carry environmental/health footprints (reproductive toxicity concerns for DMF; persistence and high COD for DMSO spills).
• Buffers: Mild aqueous buffers at pH 7.5–8.6 are standard; waste contains organic dyes that should be minimized and captured.

Greener substitutions (literature-guided)

• Solvent reduction: Prepare higher-concentration DMSO stocks (e.g., 20–50 mM) and use minimal volumes to keep final DMSO ≤1–5% v/v in reactions, reducing solvent load.
• Alternative solvents/co-solvents: Glycerol carbonate or propylene carbonate can sometimes dissolve cyanine dyes and are considered greener than DMF; verify solubility and reactivity compatibility before adoption.
• Aqueous-first approaches: Where feasible, use sulfonated Cy3 analogs (water-soluble) to avoid aprotic dipolar solvents; note that photophysics and labeling patterns can differ.
• Process changes: Micro-scale labeling and in-line desalting minimize waste. Implement solid-phase labeling on resins to improve recovery and reduce solvent volumes.

Comparative notes

• DMF vs DMSO: Prefer DMSO when possible due to lower specific hazards than DMF; ensure both are anhydrous to prevent NHS hydrolysis and repeat reactions.
• Purification: Choose gravity desalting columns over preparative RP-HPLC when purity requirements permit, cutting acetonitrile consumption.

Waste and EHS

• Segregate dye-containing waste; use activated carbon or specialized absorbents for capture before aqueous discharge. Follow local regulations for disposal of halide-containing dye salts (bromide counterion).

Pharmaceutical Uses

• Role: This product is a research-grade fluorescent labeling reagent intended for in vitro assay development, analytical method development, and process research. It is not an excipient or an approved component of drug products.
• Typical uses in pharmaceutical R&D (general):
  • Conjugation to proteins/peptides for bioanalytical method development (e.g., ligand-binding assays, imaging of process intermediates).
  • Surface labeling in biosensor platforms to support screening and characterization.
  • Spiking standards for method qualification in fluorescence-based analytics.
• Regulatory status: No pharmacopeial monograph is implied. For any use in GMP settings, assess suitability via internal qualification (identity, purity, residual solvents, counterion, bioburden/endotoxin if applicable) and reference the specific CoA/Spec Sheet.
• Documentation: Maintain batch-specific CoA, SDS, and in-house qualification reports. Define acceptance criteria for purity, residual moisture, and photophysical performance relevant to the intended application.
• Alternatives: Where GMP or IVD compliance is required, consider sourcing dyes with appropriate quality designations and documentation; specifications such as bioburden/endotoxin and residual solvent limits may be necessary.

Note: No medical or clinical claims are made. This material is for research use only, as indicated in the product data.

Physical Properties

Item-specific values

• Appearance: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular Weight: Not specified for this item; refer to CoA/Spec Sheet.
• Density, MP/BP, refractive index, elemental/metal specs: Not specified for this item; refer to CoA/Spec Sheet.
• Solubility (specification): Not specified for this item; refer to CoA/Spec Sheet.

Literature/general properties for Cy3 NHS ester–type dyes (guidance only)

• Physical state: Typically supplied as a dark-colored solid (powder or film); intensely colored, strongly light-absorbing.
• Solubility profile: Readily soluble in anhydrous polar aprotic solvents (DMSO, DMF, NMP); limited aqueous solubility unless sulfonated derivatives are used. Hydrolyzes in aqueous buffers over time (faster at higher pH).
• Spectroscopic properties (free dye, literature typical): Absorption maximum ~548–554 nm; emission maximum ~565–570 nm in aqueous/MeOH; high molar absorptivity (ε often ~1.2–1.5 × 10^5 M−1 cm−1). Exact values depend on linker, environment, and degree of aggregation.
• LogP/charge: Parent Cy3 chromophore is cationic; overall hydrophilicity is modulated by sulfonates or linkers (this item’s specific hydrophilicity is not specified).
• Stability: Moisture- and base-sensitive due to NHS ester; storage under dry, inert, light-protected conditions recommended. NHS half-life in pH 8.3 buffer is typically tens of minutes to a few hours depending on composition and temperature (literature).

Important

• Do not treat literature values as product specifications. Confirm exact values and limits for this SKU in the CoA/Spec Sheet.

Quality & Grades

Item-specific grade/purity

• Grade/Purity: Not specified for this item; refer to CoA/Spec Sheet.

Interpretation and implications (general guidance)

• Dye labeling grade: For amine-reactive dyes, quality is typically defined by dye purity (HPLC area%), identity (MS), counterion content, water content, and residual solvents. If “labeling grade” is stated in a CoA, expect low levels of hydrolyzed dye/oligomers and tight control of isomeric composition.
• HPLC-grade solvents relevance: Low-UV solvents (e.g., ACN, MeOH) are recommended for analytical QC of dye stocks to minimize background in chromatography and spectrophotometry.
• Stabilizers: Most NHS esters are supplied without stabilizers; any stabilizers or salt forms should be explicitly listed on the CoA. If stabilizers are present, assess their impact on your conjugation chemistry and downstream purification.
• Photophysical consistency: Batch-to-batch consistency is critical for quantitative labeling (e.g., DoL calculations). Review CoA for extinction coefficient reference, residual moisture, and counterion identity (bromide, as named here).
• Recommended verifications upon receipt: Record UV–Vis spectrum of a test solution (in MeOH or PBS with small DMSO) to confirm λmax profile; run analytical HPLC and/or LC–MS for identity and purity; check moisture by KF if your workflow is moisture-sensitive.

For exact specifications and acceptance criteria of SKU C1450922, refer to the product’s CoA/Spec Sheet.

Reaction & Applications

Scope (general literature/practice for Cy3 NHS esters)

• Primary function: Site-nonspecific labeling of primary amines on proteins (lysine ε-NH2, N-termini), peptides, oligonucleotides with amine modifiers, and amine-functionalized surfaces/particles to yield stable amide bonds.
• Reaction type: N-acylation via NHS ester; proceeds under mildly basic aqueous conditions with organic cosolvent.

Key applications

• Protein/antibody conjugation for fluorescence microscopy, flow cytometry, in-gel fluorescence, and immunoassays.
• Nucleic acid probe generation (5′-amine oligos) for FISH and qPCR probes.
• Bead/surface functionalization for biosensors and microarrays.

Practical tips

• Maintain anhydrous stock solutions (5–20 mM in DMSO); prepare fresh before use. Limit aqueous exposure to reduce hydrolysis.
• Typical reaction conditions (literature): pH 8.3–8.6, 20–25°C, 30–60 min for proteins; use 3–10 molar equivalents of dye vs reactive amines on the target. For antibodies, 3–6 eq per IgG often affords degree of labeling (DoL) ~2–4.
• Remove unreacted dye by gel filtration (Sephadex G-25), centrifugal desalting, or hydrophobic interaction chromatography. For oligos, use spin desalting or HPLC.
• Spectral readouts (literature typical): Cy3-labeled conjugates absorb near 548–554 nm and emit near 565–570 nm; verify spectra in your buffer to account for environment effects.

Notes

• Manufacturer Applications: Not provided in product data; the above expands on standard uses for Cy3 NHS ester reagents.
• For precise stoichiometry and extinction coefficients applicable to your system, validate empirically; do not treat these as product specifications.

Reaction Conditions

General, literature-based conditions for amine labeling with Cy3 NHS esters (not item-specific specifications):

Protein labeling

• Buffer: 50–100 mM sodium bicarbonate (pH 8.3–8.6) or phosphate (pH 7.8–8.0), amine-free.
• Concentrations: Protein 1–10 mg/mL; dye stock 5–20 mM in anhydrous DMSO. Add dye to achieve 3–10 molar equivalents per protein (or per accessible lysine for small peptides).
• Temperature/time: 20–25°C, protected from light, 30–60 min; gentle mixing.
• Workup: Quench residual activity with ethanolamine (50 mM, 10 min). Purify by desalting (G-25), spin columns, or SEC. Collect orange-red fluorescent fractions.
• Expected outcomes: Degree of labeling commonly 2–4 dyes per IgG with 3–6 eq input; verify by UV–Vis using protein A280 correction and dye Aλmax (εCy3 often ~1.2–1.5×10^5 M−1 cm−1, literature).

Oligonucleotide labeling (5′-amine)

• Solvent: 0.1 M sodium bicarbonate, pH 8.5, with 10–20% DMSO to solubilize dye.
• Stoichiometry: 5–20 eq dye vs oligo; 1–2 h at room temperature, then desalting or RP-HPLC purification.

Surface/particle labeling

• Medium: Anhydrous DMF or DMSO with catalytic base (e.g., DIPEA) for nonaqueous systems; or aqueous bicarbonate for amine-terminated hydrogels.
• Post-treatment: Thorough washing to remove physisorbed dye.

Notes

• Oxygen and light: Protect from light; oxygen has limited chemical impact on NHS coupling but affects photostability during handling.
• These conditions are typical starting points; optimize for your specific substrate and performance goals.

Safety & Handling

Item-specific hazard data

• GHS Classification: Not specified for this item; refer to SDS.
• Signal Word: Not specified for this item; refer to SDS.
• H-Statements: Not specified for this item; refer to SDS.
• Pictograms: Not specified for this item; refer to SDS.

General safety guidance for NHS-activated cyanine dyes (literature/practice)

• Likely hazards: May cause skin/eye/respiratory irritation. Dust may be harmful if inhaled. Avoid contact and inhalation. Many cyanine dyes are photosensitive and may stain.
• PPE: Lab coat, nitrile gloves (double-glove for extended handling), splash goggles. Use in a fume hood to minimize inhalation and protect from light.
• Handling: Work quickly with small quantities. Keep containers tightly closed under dry inert gas when possible. Prepare solutions in anhydrous DMSO/DMF. Avoid exposure to moisture and amine-containing vapors (NHS ester hydrolysis).
• Light sensitivity: Protect solids and solutions from ambient light (wrap vials in foil or use amber glass).
• Incompatibilities: Strong bases, nucleophiles (amines, hydrazines), water/moist air, strong oxidizers. Avoid buffers containing primary/secondary amines (e.g., Tris) during conjugation setup.
• First aid (overview; consult SDS): Eye/skin contact—rinse with water for 15 minutes, remove contaminated clothing, seek medical attention if irritation persists. Inhalation—move to fresh air; seek medical attention if symptoms occur. Ingestion—rinse mouth; seek medical advice.
• Spill/leak: Avoid dust formation. Cover with inert absorbent, collect, and dispose as hazardous chemical waste.

Always consult the product-specific SDS for authoritative safety information.

Solvent Selection

Role and polarity

• Function: Amine-reactive dye; typically dissolved as a concentrated stock in anhydrous polar aprotic solvent.
• Preferred solvents (literature/practice):
  • Anhydrous DMSO: Excellent solubility; widely used for preparing 5–20 mM stocks.
  • Anhydrous DMF or NMP: Comparable alternatives when DMSO is undesirable.
• Water miscibility: DMSO/DMF stocks can be diluted into aqueous buffers immediately before conjugation; prolonged aqueous exposure promotes NHS hydrolysis.

Buffer compatibility for conjugation (general)

• Use: Sodium bicarbonate/carbonate (pH 8.3–8.6), borate (pH 8.5–9.0), or phosphate (pH 7.4–8.0) without primary/secondary amines.
• Avoid: Tris, glycine, ethanolamine, or other nucleophile-containing buffers that quench NHS esters.

Selection tips

• Choose DMSO when highest solubility and rapid dispersion are needed; select DMF for lower viscosity or if DMSO is incompatible with biomolecules in your system.
• For highly aqueous workflows, consider sulfonated Cy3 analogs ("sulfo-Cy3") which dissolve directly in water (trade-off: altered hydrophobic interactions and potentially different labeling behavior).

Small comparison (literature)

• DMSO vs DMF: DMSO offers better dye solubilization and is less volatile; DMF provides easier removal post-reaction and lower freezing point for cold operations. Both must be anhydrous to preserve NHS activity.

Note

• No item-specific solvent specifications are provided for this SKU; the above reflects standard practice for NHS-ester cyanine dyes.

Storage & Reconstitution

Item-specific storage/shipping

• Storage Conditions: Store at -20°C (per product data). Keep vial tightly closed, desiccated, and protected from light.
• Shipped In: Ice chest + Ice pads (per product data).

General guidance for NHS-ester cyanine dyes

• Light/moisture protection: Store in amber vials or wrap in foil. Include desiccant. Consider inert gas backfill after opening.
• Reconstitution: Dissolve in anhydrous DMSO or DMF to prepare a concentrated stock (e.g., 5–20 mM). Mix gently until fully dissolved.
• Aliquoting: Immediately aliquot stock into single-use portions (low-binding microtubes), purge with inert gas if possible, and freeze at −20 to −80°C.
• Stability: Minimize freeze–thaw cycles. Avoid repeated warming; thaw just before use. Aqueous working solutions should be prepared immediately before conjugation and used within the reaction time window to limit NHS hydrolysis.
• Compatibility: Do not store dye in aqueous buffers. Avoid contact with amine-containing buffers until the intended quench step.
• Records: Note preparation date, solvent, concentration, and number of freeze–thaws for each aliquot.

Disclaimer

• Where exact shelf life or solution stability is needed, consult the product’s CoA/Spec Sheet and SDS. Research use only, as indicated.

Structure & Identity

• SKU: C1450922
• Product Name: Cy3 NHS ester bromide
• CAS: 2892098-34-7
• Category: Life science fluorescent labeling reagent (amine-reactive dye)

Item-specific identifiers

• Molecular Formula: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular Weight: Not specified for this item; refer to CoA/Spec Sheet.
• SMILES: Not specified for this item; refer to CoA/Spec Sheet.
• InChIKey: Not specified for this item; refer to CoA/Spec Sheet.

General structural description (literature, representative of Cy3 NHS esters)

• Core scaffold: An indocarbocyanine (Cy3) chromophore consisting of two indolenine/benzindolenine rings linked by a trimethine bridge (polymethine chain), conferring strong absorption in the green-yellow and emission in the orange-red.
• Charge/counterion: Cy3 dyes are typically cationic; “bromide” denotes a bromide counterion balancing the dye’s positive charge (literature).
• Reactive handle: An N-hydroxysuccinimide (NHS) active ester linked via a carboxyl-bearing spacer to the chromophore; reacts selectively with primary amines to form stable amide bonds (literature).
• 2D description: Two quaternized indoleninium rings at the termini, conjugated through a –CH=CH–CH= chain; a pendant linker terminating in an NHS succinimide ring (five-membered imide) serves as the acylating group. No defined stereocenters on the chromophore; E/Z isomerism may exist along the polymethine chain in solution (literature).

Notes

• Exact substitution pattern, linker length, and counterion stoichiometry can vary across commercial Cy3 NHS derivatives; for this specific catalog item, consult the CoA/Spec Sheet for definitive structural identifiers.

Synthetic Utility

While primarily a bioconjugation reagent, Cy3 NHS ester bromide also has defined chemical reactivity valuable to synthetic and materials chemists (general literature guidance):

• Functional group: Activated carboxylic acid derivative (NHS ester). Reacts with primary amines to form amide bonds; secondary amines react more slowly; tertiary amines do not acylate.
• Selectivity: In mixed-nucleophile environments, primary aliphatic amines are favored over phenols, thiols (at neutral pH), and imidazoles. Thiols can react under basic, nonaqueous conditions but are not preferred for NHS esters.
• Surfaces/materials: Enables covalent grafting of the Cy3 chromophore to amine-terminated polymers, silica, nanoparticles, and microarray substrates, allowing optical encoding and sensing.
• Retrosynthetic value: The NHS ester masks a carboxylate, providing a shelf-stable (when dry) activated intermediate compared to acid chlorides. It can be coupled under mild, nearly neutral conditions that are compatible with delicate substrates.
• Orthogonality: Compatible with many protecting groups and click handles; can be integrated into multi-step labeling sequences where amine coupling precedes/ follows azide–alkyne cycloaddition or thiol–maleimide chemistry (ensure sequence avoids NHS hydrolysis).
• Photophysical handle: The chromophore’s strong absorbance enables stoichiometric monitoring of coupling progress by UV–Vis.

Caveats

• Hydrolysis competes with coupling in aqueous media; optimize pH, time, and dye concentration.
• The bromide counterion is generally a spectator; however, ionic strength may influence aggregation and surface interactions.

Target Specificity

This product is a general amine-reactive fluorescent labeling reagent and does not possess intrinsic biological target specificity (no antigen/epitope/clonal attributes). It reacts chemically with accessible primary amines on diverse substrates.

Item-specific targeting data

• Antigen/epitope: Not applicable.
• Species reactivity: Not applicable.
• Clone/isotype: Not applicable.

Notes

• Selectivity is governed by chemistry (amine availability and pKa), not by biological recognition. For site-specific biomolecular conjugation, consider orthogonal strategies (e.g., engineered cysteine/maleimide, enzymatic tags, click chemistry).