Bis-sulfone-PEG4-NHS Ester - ≥93% , CAS No.2055047-19-1

CAS: 2055047-19-1 Cat. No.: B597116 Formula: C₄₀H₄₈N₂O₁₄S₂ Peso molecolare: 844.94

GRADE & PURITY ≥93%

Synonyms

(2,5-dioxopyrrolidin-1-yl) 3-[2-[2-[2-[2-[[4-[3-(4-methylphenyl)sulfonyl-2-[(4-methylphenyl)sulfonylmethyl]propanoyl]benzoyl]amino]ethoxy]ethoxy]ethoxy]ethoxy]propanoate

Storage

Store at -20°C,Argon charged,Desiccated

Shipped In

Ice chest + Ice pads

Application

Antibody Drug Conjugates(ADC), Drug Delivery, Surface Modification, PEGylation of Protein, Peptide & Oligo

Panoramica

Bis-sulfone-PEG4-NHS Ester is a bis-alkylating labeling reagent that is selective for the cysteine sulfur atoms from a native disulfide. These reagents undergo bis-alkylation to conjugate both thiols derived from the two cysteine residues of a reduced native disulfide bond such as the interchain disulfide bonds of an antibody. The reaction results in covalent rebridging of the disulfide bond via a three carbon bridge leaving the protein structurally intact. The hydrophilic PEG spacer increases solubility in aqueous media.

Specifications

Sinonimi

(2,5-dioxopyrrolidin-1-yl) 3-[2-[2-[2-[2-[[4-[3-(4-methylphenyl)sulfonyl-2-[(4-methylphenyl)sulfonylmethyl]propanoyl]benzoyl]amino]ethoxy]ethoxy]ethoxy]ethoxy]propanoate

Specifiche e purezza

≥93%

Condizioni di conservazione di stoccaggio

Store at -20°C,Argon charged,Desiccated

Spedito in

Ice chest + Ice pads

Questo prodotto richiede spedizione a catena fredda. I servizi di terra e altri servizi economici non sono disponibili.

Purezza

≥93%

Nomi e identificatori

• Peso molecolare: 844.94

Proprietà chimiche e fisiche

• Solubilità: DMSO, DCM, DMF
• Sensibilità: Moisture sensitive;Heat sensitive

Application Protocols

Illustrative protocol for amine coupling followed by cysteine rebridging (literature; optimize per system)

1. Preparation

• Equilibrate protein (e.g., antibody) in amine-free buffer (PBS or HEPES, pH 7.4–7.8). Concentration: 1–10 mg/mL.
• Prepare fresh, anhydrous stock of Bis-sulfone-PEG4-NHS Ester in DMSO (10–50 mM). Keep on ice and protected from moisture.

2. NHS–amine coupling

• Add linker stock to protein to reach 5–10 molar eq per protein (or per target amine site if known). Keep final DMSO ≤10% v/v.
• Incubate 20–25 °C for 30–60 min with gentle mixing.
• Quench unreacted NHS with 20–50 mM ethanolamine, 10 min at RT.
• Desalt (G-25) or perform UF/DF to remove small molecules.

3. Disulfide reduction (if rebridging intended)

• Treat with TCEP (0.5–2 mM) at pH 7.0–7.5 for 15–30 min at RT. Remove excess TCEP if required by downstream analytics.

4. Bis-sulfone–thiol conjugation/rebridging

• Adjust to pH 6.8–7.4. Incubate 30–120 min at RT to allow thiol additions to the bis-sulfone handle.
• Monitor by non-reducing SDS-PAGE and intact MS.

5. Final cleanup and characterization

• Purify by SEC or UF/DF. Quantify conjugation (DAR/DOL) by UV–Vis or MS. Confirm absence of free thiols (Ellman’s assay).

Notes

• Avoid Tris/glycine buffers during NHS coupling. Exclude competing thiols during the bis-sulfone step. Conduct small-scale pilot reactions to optimize equivalents and timing.

These steps are general literature guidance and are not validated protocols for this specific item.

Biological Roles

This product is a synthetic chemical linker for research use and does not possess an intrinsic biological role.

General biochemical context (literature; not product-specific claims)

• Conjugation chemistry: NHS esters target primary amines (lysine residues, N-termini) to form amide bonds, a reaction commonly exploited to attach functional handles to biomolecules.
• Cysteine rebridging: Bis-sulfone motifs are engineered electrophiles that can form bridges between two thiols originating from a reduced disulfide, stabilizing protein tertiary structure while introducing a defined modification site.
• PEG4 spacer: Short PEG segments can improve aqueous compatibility and reduce nonspecific interactions in biomolecular environments. PEGylation often decreases protein adsorption to surfaces and can modulate hydrodynamic properties of conjugates.
• Applications in assay development: Such linkers enable creation of enzyme–inhibitor, antibody–reporter, or ligand–biomaterial conjugates for analytical, diagnostic research, and materials science platforms.

Note

• No therapeutic, diagnostic, or clinical utility is asserted. Use is restricted to laboratory research by qualified personnel.

Buffer Applications

This compound is not a buffer and is not intended to establish or maintain solution pH.

Practical buffer guidance for using NHS-ester linkers (literature)

• Suitable buffers for NHS coupling: Phosphate (10–100 mM, pH 7.2–7.8), HEPES (pH 7.2–8.0), or carbonate/bicarbonate (pH 8.3–8.5) depending on desired rate/selectivity. Maintain ionic strength (100–150 mM NaCl) for protein stability when appropriate.
• Avoid primary amine buffers: Tris, glycine, and ammonium salts compete with amine targets, quenching the NHS ester.
• Additives: 0.01–0.05% nonionic surfactant (Tween-20) can reduce aggregation for sensitive proteins. EDTA (0.5–1 mM) can chelate adventitious metals that catalyze oxidation during thiol steps.
• For thiol conjugation (bis-sulfone step): Use neutral buffers (PBS, pH 6.8–7.4) to favor thiol addition while minimizing amine addition; ensure absence of thiol reducing agents (DTT, β-ME) that would compete unless used catalytically and removed prior to reaction.
• Handling: Prepare fresh buffer, degas if necessary, and filter (0.2 µm) to remove particulates for bioconjugation workflows.

Green Alternatives

Context (literature; general sustainability considerations)

• Challenge: NHS-ester linkers are inherently moisture-sensitive and commonly require polar aprotic solvents (DMSO, DMF, NMP) with recognized environmental/health footprints. Greener choices aim to minimize or replace these while preserving activity.

Comparative options for handling and solvent systems (informational)

• DMSO (benchmark): Widely accepted in bioconjugation; relatively benign compared with DMF/NMP but persistent in aqueous waste. Low volatility reduces inhalation exposure.
• DMF/NMP: Effective but subject to stricter regulatory scrutiny. Consider replacing with:
  • PC/GBL (propylene carbonate, γ-butyrolactone): High-permittivity, low vapor pressure; can dissolve PEGylated linkers. Verify compatibility with biomolecules and downstream assays.
  • Cyrene (dihydrolevoglucosenone): Bio-based dipolar aprotic; sometimes suitable for linker dissolution, but viscosity and reactivity must be assessed; limited bioconjugation precedent.
  • Aqueous-organic microvolumes: Pre-dissolve in minimal DMSO, then rapidly dilute into buffered saline immediately before use to minimize aprotic solvent content.

Process improvements

• Reduce solvent: Use concentrated stocks (50–100 mM) and minimal dosing volumes to lower total solvent footprint.
• Single-use aliquots: Prevents degradation and waste from repeated thawing.
• Room-temperature operations: Avoid heating to reduce energy input and side reactions.

Trade-offs

• Greener solvents may alter solubility and reaction kinetics; perform pilot solubility and reactivity tests. Maintain dry conditions to preserve NHS activity regardless of solvent choice.

Pharmaceutical Uses

No clinical or therapeutic claims are made for this product. It is supplied strictly for research use only.

Formulation/manufacturing context (literature; general)

• Role: Bis-sulfone-PEG4-NHS ester functions as a heterobifunctional crosslinker for constructing well-defined conjugates (e.g., model antibody–linker–payload systems) during discovery-stage process development.
• Advantages: PEG4 spacing can reduce hydrophobicity of conjugates, improving solubility and potentially lowering aggregation during formulation screening relative to purely hydrophobic linkers.
• Process considerations: During early development, linker stocks are prepared in sterile, anhydrous DMSO or DMF and added to protein formulations at controlled solvent percentages (commonly ≤10% v/v before further dilution). Post-conjugation, buffer exchange, ultrafiltration/diafiltration (UF/DF), or SEC can remove residual small molecules.
• Analytical controls: Monitor degree of labeling (DOL) or drug-to-antibody ratio (DAR) by intact mass spectrometry, UV/Vis deconvolution, or hydrophobic interaction chromatography (HIC). Assess free thiols (Ellman’s assay) before and after rebridging.
• Regulatory note: For any translational work, material grade, residual solvents, extractables/leachables, and impurity profiles must be established separately. This listing provides no pharmacopeial status or GMP assurance.

Physical Properties

Item-specific (from Product Data)

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

Literature/general expectations for this class of reagent (informational only; not product specifications)

• Physical state: Typically a white to off-white solid or foam for NHS-ester PEG linkers; hygroscopic.
• Solubility (qualitative): High in polar aprotic solvents (DMSO, DMF, NMP); moderate in acetonitrile; limited in water until hydrolyzed; insoluble in nonpolar hydrocarbons.
• Hydrolytic behavior: NHS esters hydrolyze in the presence of moisture, faster at higher pH and temperature; half-life can range from minutes (pH > 8.5) to hours (pH ~7.0) depending on solvent and buffer.
• Partitioning: PEG4 imparts hydrophilicity, typically lowering logP relative to purely hydrophobic linkers, improving dispersion in aqueous-organic mixtures.
• UV characteristics: NHS and PEG segments have weak chromophores; bis-sulfone may exhibit modest absorbance in the near-UV; for analytical monitoring, LC–MS or derivatization-based methods are often preferred.
• Thermal behavior: Many NHS ester PEGylated linkers soften below 100 °C and decompose before a sharp melting point; avoid elevated temperatures to limit hydrolysis and decomposition.

Not provided for this specific item (consult CoA/Spec Sheet for confirmed values)

• Melting/boiling point, density, refractive index, logP, pKa, water content, residual solvents, metals, UV cutoff.

Quality and Grades

Item-specific (from Product Data)

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

Guidance on interpreting quality for this reagent class (general)

• Purity reporting: For functional crosslinkers, both assay purity and functional activity (residual NHS content) are relevant. Vendors may report HPLC area %, water content (KF), residual solvents, and identity by NMR/HRMS.
• Functional integrity: NHS ester content diminishes with moisture exposure; recent CoA date and sealed, desiccated handling are indicators of maintained activity.
• Stabilizers: These materials are typically supplied without stabilizers; adding bases, amines, or aqueous buffers is contraindicated as it promotes hydrolysis. If provided with desiccant or under inert gas, retain original packaging conditions.
• Chromatographic grade vs. bioconjugation grade: “Bioconjugation” or “Protein labeling” grade often implies low bioburden, low residual acids/bases, and high functional group integrity suitable for sensitive biomolecule work. Absent a declared grade, verify suitability with small-scale pilot reactions.
• Documentation: For regulated workflows, request and review CoA/Spec Sheet for batch-specific specifications, analytical methods, and recommended handling limits (water, peroxide, metals if applicable).

Reaction and Applications

General applications (literature; expand-on context, not product-specific validation)

• Heterobifunctional conjugation: The NHS ester acylates primary amines on peptides, proteins, or surface-bound amino groups to install a bis-sulfone handle via a stable amide linkage.
• Cysteine targeting via bis-sulfone: The bis-sulfone acts as a double Michael acceptor, enabling selective conjugation to thiols. In proteins, reduced disulfide pairs can be “rebridged,” producing a stable, site-constrained thioether bridge that maintains protein architecture better than random alkylation.
• Antibody–drug conjugate (ADC) methodology: After mild reduction of interchain disulfides (e.g., TCEP), bis-sulfone reagents can install payloads or handles across cysteine pairs, improving DAR control and conjugate homogeneity compared to stochastic lysine labeling.
• Surface/biomaterial functionalization: The NHS end enables immobilization onto amine-rich polymers (e.g., PEI, chitosan, aminated glass), placing a thiol-reactive bis-sulfone moiety for subsequent bioconjugation.
• Multistep orthogonal assembly: Sequence amine coupling (NHS) then thiol conjugation (bis-sulfone) to assemble complex architectures (protein–polymer, protein–protein, or protein–small molecule).

Practical tips

• Maintain pH 7.4–8.5 for NHS coupling (10–60 min at RT) and pH 6.8–7.8 for thiol additions (to limit undesired amine addition). Use reducing agents like TCEP (non-thiol) when generating free cysteines; avoid DTT/β-ME in the thiol-conjugation step as they compete.
• Control stoichiometry: 2–10 eq linker per lysine site for amine coupling; 1–2 eq per reduced disulfide for rebridging. Optimize by small-scale trials.
• Quench unreacted NHS esters with ethanolamine or Tris after amine coupling, prior to thiol chemistry if needed.

Reaction Conditions

General guidance (literature; adjust per substrate)

• NHS–amine coupling:
  • Solvent/buffer: PBS or HEPES (pH 7.4–7.8) or carbonate (pH 8.3–8.5). Pre-dissolve reagent in dry DMSO/DMF (10–50 mM), add to achieve ≤10% organic v/v.
  • Stoichiometry: 2–20 eq linker per amine target on proteins; 1.1–2.0 eq for small molecules.
  • Temperature/time: 20–25 °C for 15–60 min (proteins) or up to 2 h (small molecules). Monitor by LC–MS or ninhydrin loss of amines.
  • Quench: Ethanolamine or Tris (10–50 mM) to consume residual NHS after desired coupling.
• Thiol addition/rebridging (bis-sulfone step):
  • Generation of thiols: Reduce disulfides with TCEP (0.5–5 mM) at pH 7.0–7.5; remove excess TCEP if it interferes with downstream analytics (it is non-thiol and generally compatible).
  • Reaction: pH 6.8–7.6 in PBS; 20–25 °C; 30–120 min. Use 1–2 eq per disulfide for rebridging. Avoid competing thiols (DTT, β-ME).
  • Verification: Non-reducing SDS-PAGE for rebridging, intact MS for mass shift, Ellman’s assay to confirm consumption of free thiols.
• Workup/purification:
  • Proteins: Desalting (G-25), dialysis, or UF/DF (10–50 kDa MWCO). Small molecules: silica or reverse-phase HPLC.
• Stability: Perform reactions promptly after preparing solutions; NHS hydrolysis accelerates at elevated pH and temperature.

All conditions above are literature best practices and not specifications for this particular item.

Safety and Handling

Item-specific (from Product Data)

• GHS Classification / Signal Word / H-Statements / Pictograms: Not specified for this item; refer to SDS.
• Storage Conditions: Store at -20°C.
• Shipped In: Ice chest + Ice pads.

General safety guidance for NHS-ester/bis-sulfone linkers (literature; defer to SDS)

• Hazards: NHS esters are acylating agents that can cause skin/eye irritation. Bis-sulfone Michael acceptors may alkylate thiols. Avoid inhalation of dust and contact with skin/eyes.
• PPE: Laboratory coat, safety glasses, and appropriate chemical-resistant gloves (e.g., nitrile). Handle in a fume hood to control dust and solvent vapors.
• Incompatibilities: Moisture, bases, and nucleophiles (amines, thiols) accelerate decomposition. Avoid primary-amine buffers (e.g., Tris, glycine) during handling/conjugation of the NHS ester stage.
• First aid overview: In case of skin/eye contact, rinse with water for ≥15 minutes and seek medical attention. If inhaled, move to fresh air. If ingested, do not induce vomiting; seek medical advice. Always follow the SDS.
• Spill/cleanup: Avoid dust generation. Collect solids with inert absorbent, place in suitable container for disposal per institutional guidelines.
• Stability notes: Keep container tightly closed under dry, inert atmosphere. Minimize freeze–thaw cycles and exposure to ambient humidity to limit NHS hydrolysis.

Disposal

• Dispose of unused material and contaminated consumables as hazardous chemical waste per local regulations and institutional EHS policies.

Solvent Selection

General guidance (literature; not product specifications)

• Polarity/miscibility: The PEG4 linker imparts substantial polarity and hydrogen-bond acceptor capacity, making the reagent readily soluble in polar aprotic solvents (DMSO, DMF, NMP) and miscible with water when pre-dissolved in these solvents.
• Recommended solvents for stock solutions: Anhydrous DMSO or DMF (typical 10–50 mM stocks). Filter-sterilize through 0.2 µm if using in bioconjugation.
• Aqueous handling: NHS esters hydrolyze in water; if aqueous use is required, dilute the anhydrous stock into buffer immediately before use and consume within the hydrolytic half-life window.
• Buffer compatibility during NHS coupling: Avoid primary amine buffers (Tris, glycine, ammonium bicarbonate). Use phosphate, HEPES, or carbonate buffers. Maintain pH 7.4–8.5 to balance reaction rate vs. hydrolysis.
• Removal of solvents: Post-reaction, excess linker can be removed by desalting columns (e.g., Sephadex G-25), spin columns, ultrafiltration (10–50 kDa MWCO for proteins), or preparative chromatography, depending on application.
• Comparison to alternatives:
  • DMSO vs. DMF: DMSO offers superior biocompatibility upon dilution but can participate weakly in nucleophilic exchanges at high temperature; DMF provides similar solubilization with lower viscosity.
  • ACN/IPA: Often inadequate for full dissolution; may be used as minor co-solvents to adjust viscosity/volatility.
• Drying: Use freshly opened anhydrous solvent or dry over molecular sieves (3Å/4Å) to maximize NHS integrity.

Storage and Reconstitution

Item-specific (from Product Data)

• Storage Conditions: Store at -20°C.
• Shipped In: Ice chest + Ice pads.
• Research Use Note: For research use only.

General handling recommendations (literature; not product specifications)

• Moisture sensitivity: NHS esters hydrolyze; store desiccated under inert gas (argon/nitrogen) when possible. Keep container tightly closed.
• Light/temperature: Protect from light and avoid repeated warming. Short handling at room temperature is acceptable if dry.
• Reconstitution: Dissolve in anhydrous DMSO or DMF to 10–50 mM. Vortex briefly and, if required, sonicate gently. Use 0.2 µm PTFE filtration for sterile work.
• Aliquoting: Prepare single-use aliquots immediately after opening to minimize freeze–thaw and headspace moisture. Freeze aliquots at -20 to -80 °C in moisture-tight vials.
• In-use stability: Once diluted into aqueous buffer, consume promptly (within the expected NHS half-life at the working pH). Keep samples on ice during setup to slow hydrolysis.
• Disposal of residues: Quench small amounts by hydrolysis (aqueous buffer) before disposal per institutional guidelines.

Not specified for this item (consult CoA/Spec Sheet)

• Exact shelf-life/retest date, water content limits, recommended desiccant type, or stabilizers (if any).

Structure and Identity

Item-specific (from Product Data)

• SKU: B597116
• Product Name: Bis-sulfone-PEG4-NHS Ester
• CAS: 2055047-19-1
• Molecular Formula: Not specified for this item; refer to CoA/Spec Sheet.
• Molecular Weight: Not specified for this item; refer to CoA/Spec Sheet.
• InChIKey: Not specified for this item; refer to CoA/Spec Sheet.
• SMILES: Not specified for this item; refer to CoA/Spec Sheet.

General structural description (literature/context)

• Functional architecture: A heterobifunctional linker comprising:
  • An NHS (N-hydroxysuccinimide) activated ester for rapid acylation of primary amines (e.g., lysine side chains, N-termini) to form stable amide bonds.
  • A “bis-sulfone” motif, typically functioning as a double Michael acceptor toward thiols, enabling conjugation to two cysteines (e.g., disulfide rebridging) to afford a stable thioether bridge.
  • A PEG4 spacer (four ethylene glycol units) providing hydrophilicity, reduced aggregation, and improved aqueous handling while distancing reactive groups to reduce steric hindrance.
• 2D features (described): Succinimide ring (five-membered imide) attached to an activated carbonate/carboxyl center; a flexible –(O–CH2–CH2)4– segment; and a bis-activated sulfone-containing Michael acceptor segment capable of sequential thiol additions.
• Stereochemistry: Typically none specified; PEG and NHS modules are achiral. Any chiral centers would be uncommon and are not implied by the name.

Notes

• Exact regiochemistry, terminal groups, and protecting groups can vary across vendors; consult the item’s CoA/Spec Sheet for definitive structure and elemental data.

Synthetic Utility

General chemical reactivity (literature)

• NHS ester: Activated carboxyl derivative that undergoes nucleophilic acyl substitution with primary amines to form amides. Reaction rate increases with pH (7.4–8.5), but hydrolysis competes; anhydrous organic cosolvents and controlled buffering improve selectivity.
• Bis-sulfone electrophile: Functions as a double Michael acceptor, capable of sequential thiol additions. In the context of reduced disulfide bonds, it can rebridge two cysteines, yielding a stable thioether linkage and restoring covalent connectivity.
• PEG4 spacer: Enhances solubility and can modulate spacing to reduce steric hindrance in surface and biomolecule conjugations.

Use cases in synthesis and materials

• Stepwise orthogonal assembly: Install NHS-derived amide on an amine-bearing scaffold, then capture thiol-bearing partners using the bis-sulfone handle.
• Surface patterning: Aminated substrates (silica, polymer films) can be functionalized to present thiol-reactive sites for immobilization of peptides/proteins with engineered cysteines.
• Modular payload attachment: Combine with thiol-bearing drugs/fluorophores or with cysteine-rich peptides to generate defined bioconjugates.

Tips and caveats

• Control pH and moisture to minimize NHS hydrolysis; prepare small, fresh aliquots.
• Avoid thiol-containing additives during the bis-sulfone step unless they are the intended reactants.
• Purification: Use preparative HPLC or size-exclusion for small-molecule vs. macromolecule separations, respectively.

Target Specificity

Not applicable to this product. This is a chemical linker, not a biological targeting reagent (no antigen/epitope, clone, isotype, or species specificity). Targeting arises from reactive group chemistry (amines and thiols), not from molecular recognition.