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Sulfo-SBED Biotin Label Transfer Kit - Western Blot Application Thermo Scientific™

The Thermo Scientific Pierce Sulfo-SBED Biotin Label Transfer Reagent is a multifunctional reagent for labeling a purified protein and then covalently transferring the attached biotin tag onto specific interactors of that protein.

Sulfo-SBED is the abbreviation for Sulfo-N-hydroxysuccinimidyl-2-(6-[biotinamido]-2-(p-azido benzamido)-hexanoamido) ethyl-1,3'-dithioproprionate. It is a heterobifunctional chemical crosslinker capable of covalently attaching to primary amines at one end and to nearly any protein functional group at the other end. Unlike typical crosslinkers, Sulfo-SBED also includes a biotin group and a cleavable disulfide spacer arm. Together these features allow one to sequentially crosslink interacting proteins and transfer the biotin affinity tag from one protein (i.e., a purified 'bait' protein) to another (possibly unknown 'prey' protein). Label Transfer is a powerful in vitro method for protein interaction discovery. A growing number of publications feature the use of Sulfo-SBED Biotin Label Transfer Reagent to identify previously unknown protein interaction binding partners and to more fully characterize the specific protein binding domains of other protein interactions.

Typical protocol for label transfer experiment:

• Add a few microliters of dissolved Sulfo-SBED Reagent to 0.5-1 mL of purified bait protein in PBS.
• Incubate mixture for 30-120 minutes on ice or at room temperature in the dark.
• Desalt or dialyze (in subdued light) to remove excess non-reacted Sulfo-SBED from the labeled bait protein.
• Add labeled bait protein to cell lysate or other solution containing putative target protein interactors ('prey').
• When interaction complexes have formed, expose the solution to ultraviolet light (365 nm) for several minutes.
• Analyze products by one of several methods:
Western Blotting: Cleave crosslinks in DTT, separate proteins by SDS-PAGE, and detect biotinylated bands by Western blotting with streptavidin-HRP.
Purification and Mass Spec or Sequencing: Affinity-purify biotinylated proteins or peptide fragments following trypsin digestion and perform MS or sequencing to characterize the proteins involved.

Related Products
Sulfo-SBED Biotin Label Transfer Reagent

Pierce™ Premium Grade EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride) Thermo Scientific™

Thermo Scientific Pierce Premium Grade EDC is our highest quality formulation of this popular carbodiimide crosslinker, specially characterized for applications where product integrity and risk minimization are paramount.

Features of Premium Grade EDC:

EDC—carbodiimide crosslinker for conjugation between carboxylates and amines
High quality—identity and purity confirmed by several tests
Product integrity—enhanced level of testing and characterization compared to standard grade
Lot retention—ample supply of past lots retained to ensure future process testing
Change management—Change Control Notification (CCN) service
Consistent manufacture—batch-specific manufacturing documentation review

Compared to the standard grade product, Premium Grade EDC provides more clearly defined quality and product support by including (a) increased analytical testing and product characterization, (b) greater batch-specific information and quality assurance review, (c) extensive lot sample retention and (d) change control notification. EDC is 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride, a water-soluble carbodiimide crosslinker that activates carboxyl groups for spontaneous reaction with primary amines, enabling peptide immobilization and hapten-carrier protein conjugation. In the presence of N-hydroxysulfosuccinimide (sulfo-NHS), EDC can be used to convert carboxyl groups to amine-reactive sulfo-NHS esters. This is accomplished by mixing the EDC with a carboxyl containing molecule and adding sulfo-NHS.

Specifications of Premium Grade EDC
We manufacture crosslinking reagents to ensure the highest possible overall product integrity, consistency and performance for the intended research applications.
• Visual: White crystalline powder, free of foreign matter
• Identity: IR scan shows only peaks characteristic of EDC
• Water content: Karl Fischer titration, ≤ 2%
• Trace metals: All metals, including lead, ≤ 50 ppm
• Elemental analysis: Reported values for C, H, and N

Applications:
• Conjugate carboxyl and amino groups among peptides and proteins
• Couple haptens to immunogenic carrier proteins (e.g., attach a peptide to KLH)
Immobilize peptide antigens to affinity-purify antibodies
• Create NHS-activated, amine-reactive labeling compounds
• Crosslink proteins to carboxyl coated beads or surfaces
• Activate nanoparticles with amine-reactive sulfo-NHS esters

Product References:
Crosslinker Application Guide -- search for recent literature references for this product

Related Products
EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride)
Pierce™ Premium Grade Sulfo-NHS (N-hydroxysulfosuccinimide)

BM(PEG)3 (1,11-bismaleimido-triethyleneglycol) Thermo Scientific™

Thermo Scientific Pierce BM(PEG)3 is a bismaleimide-activated PEG compound for crosslinking between sulfhydryl (—SH) groups in proteins and other thiol molecules. The maleimide groups at either end of the PEG3 spacer react specifically and efficiently with reduced sulfhydryls at pH 6.5-7.5 to form stable thioether bonds.

Features of BM(PEG)3:

Reactive groups: maleimide (both ends)
Reactive towards: sulfhydryl groups
• Long, pegylated, sulfhydryl-to-sulfhydryl crosslinkers, composed of maleimide groups and 3-unit polyethylene glycol spacer arm
• PEG spacers provide unique advantages, including enhanced solubility, increased stability, reduced tendency toward aggregation and reduced immunogenicity
• Pure compounds with defined structure and molecular weight, ensuring reproducible protein-modification effects
• Ideal for small molecule or peptide conjugations

Alternative Names for BM(PEG)n:

• Bismaleimide-dPEG
• Mal-PEG-Mal
• BM(PEO)2
• BM-PEO3

Product References:

Crosslinker Application Guide -- search for recent literature references for this product

Related Products
BMB (1,4-bismaleimidobutane)
BMH (bismaleimidohexane)
BMOE (bismaleimidoethane)
BM(PEG)2 (1,8-bismaleimido-diethyleneglycol)

EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride) Thermo Scientific™

Thermo Scientific Pierce EDC is a carboxyl- and amine-reactive zero-length crosslinker. EDC reacts with a carboxyl group first and forms an amine-reactive O-acylisourea intermediate that quickly reacts with an amino group to form an amide bond with release of an isourea by-product. The intermediate is unstable in aqueous solutions and so two-step conjugation procedures rely on N-hydroxysuccinimide (NHS) for stabilization. Failure to react with an amine will result in hydrolysis of the intermediate, regeneration of the carboxyl, and release of an N-substituted urea. A side reaction is the formation of an N-acylurea, which is usually restricted to carboxyls located in hydrophobic regions of proteins.

Thermo Scientific No-Weigh products are specialty reagents provided in a pre-aliquoted format. The pre-weighed packaging prevents the loss of reagent reactivity and contamination over time by eliminating the repetitive opening and closing of the vial. The format enables use of a fresh vial of reagent each time, eliminating the hassle of weighing small amounts of reagents and reducing concerns over reagent stability.

Characteristics of EDC:

Reactive group: carbodiimide
Reaction target: activates carboxyl groups to conjugate to amino groups (primary amines)
Several conjugation strategies—react EDC alone with target groups or include NHS or Sulfo-NHS to increase reaction efficiency or to stabilize active intermediate for later reaction to amines
Neutral linkage—forms neutral amide bonds between carboxyls and amines
Water-soluble reagent—add directly to reactions in aqueous, physiological buffers
Soluble reaction byproducts—easily removed by washing with water or dilute acid
High-purity, crystalline reagent—use to create high-quality, activated derivatives

Properties of EDC
• Molecular formula: C8H17N3·HCl
• Molecular weight: 191.7
• Spacer arm length: 0 Å
• CAS Number: 25952-53-8
• Reactive groups: carbodiimide
• Reactivity: Forms active intermediate with carboxyl groups at pH 4.7–6.0 (optimum), then intermediate reacts with primary amines

1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC or EDAC) is a zero-length crosslinking agent used to couple carboxyl groups to primary amines. This crosslinker has been used in diverse applications such as forming amide bonds in peptide synthesis, attaching haptens to carrier proteins to form immunogens, labeling nucleic acids through 5' phosphate groups and creating amine-reactive NHS-esters of biomolecules. EDC reacts with a carboxyl to form an amine-reactive O-acylisourea intermediate. If this intermediate does not encounter an amine, it will hydrolyze and regenerate the carboxyl group. In the presence of N-hydroxysulfosuccinimide (Sulfo-NHS), EDC can be used to convert carboxyl groups to amine-reactive Sulfo-NHS esters. This is accomplished by mixing the EDC with a carboxyl containing molecule and adding Sulfo-NHS.

Applications:
• Conjugate carboxyl and amino groups among peptides and proteins
• Couple haptens to immunogenic carrier proteins (e.g., attach a peptide to KLH)
Immobilize peptide antigens to affinity-purify antibodies
• Create NHS-activated, amine-reactive labeling compounds
• Crosslink proteins to carboxyl coated beads or surfaces
• Activate nanoparticles with amine-reactive Sulfo-NHS esters
• DNA labeling through 5' phosphate groups (see Tech Tip #30)

Product References:
Crosslinker Application Guide -- search for recent literature references for this product

BS3 (bis(sulfosuccinimidyl)suberate) Thermo Scientific™

Thermo Scientific Pierce BS3 (Sulfo-DSS) is bis(sulfosuccinimidyl)suberate, an amine-to-amine crosslinker that is homobifunctional, water-soluble, non-cleavable and membrane impermeable.

BS3 contains an amine-reactive N-hydroxysulfosuccinimide (NHS) ester at each end of an 8-carbon spacer arm. NHS esters react with primary amines at pH 7-9 to form stable amide bonds, along with release of the N-hydroxysulfosuccinimide leaving group. Proteins, including antibodies, generally have several primary amines in the side chain of lysine (K) residues and the N-terminus of each polypeptide that are available as targets for NHS-ester crosslinking reagents.

No-Weigh microtube handling:
Each of the eight microtubes contains 2 mg of crosslinker. Puncture foil with a pipette tip and add DMSO or DMF to the DSS or aqueous buffer to the BS3. Store the microtube strip in the foil pouch provided. Used microtubes may be cut from the unused microtubes and discarded.

Features of BS3:

Reactive groups: sulfo-NHS ester (both ends)
Reactive towards: amino groups (primary amines)
• Amine-reactive Sulfo-NHS ester reacts rapidly with any primary amine-containing molecule
• Water-soluble; compare with DSS
• Membrane-impermeable, allowing for cell surface labeling
• High-purity, crystalline reagent can be used to create high-purity crosslinked conjugates

Because it contains the hydrophilic sulfonyl moiety, BS3 crosslinker is soluble up to ~100 mM in water and many commonly used buffers, thus avoiding the use of organic solvents which may perturb protein structure. DSS, the non-water soluble analog of BS3 is also available for applications that require a less hydrophilic crosslinker (e.g., to effect intracellular crosslinking). DSS and BS3 have essentially identical crosslinking activity toward primary amines.

Properties of BS3:
• Alternative names: Sulfo-DSS
• Molecular formula: C16H18N2O14S2Na2
• Molecular weight: 572.43
• Spacer arm length: 11.4 Å (8 atoms)
• CAS number: 82436-77-9
• Reactive groups: NHS esters, react with primary amines at pH 7.0-9.0

Applications:
• Crosslink cell-surface proteins prior to cell lysis and immunoprecipitation
• Identify receptor-ligand interactions by chemical crosslinking
• 'Fix' protein interactions to allow identification of weak or transient protein interactions
• Identify near-neighbor protein interactions
• Protein crosslinking to create bioconjugates via single-step reactions
• Immobilize proteins onto amine-coated surfaces

BS3 Specifications:
We manufacture BS3 to the highest specifications to produce the most specific bioconjugates, ensure the integrity of your data and to provide you with the highest degree of consistency. Each lot of BS3 is tested to meet the following minimum specifications:
• Purity: > 93% by quantitative NMR (the highest standard for crosslinker purity)
• Solubility: > 5.8 mg/mL in DI water, clear solution with no insoluble material

Product References
Crosslinker Application Guide -- search for recent literature references for this product

Related Products
Pierce™ Premium Grade BS3
DSS (disuccinimidyl suberate)
BS(PEG)5 (PEGylated bis(sulfosuccinimidyl)suberate)
BS(PEG)9 (PEGylated bis(sulfosuccinimidyl)suberate)

SBAP (succinimidyl 3-(bromoacetamido)propionate) Thermo Scientific™

Thermo Scientific Pierce SBAP is a short (6.2 angstrom) crosslinker for amine-to-sulfhydryl conjugation via N-hydroxysuccinimide (NHS) ester and bromoacetyl reactive groups.

Features of SBAP:

Reactive groups: NHS ester and bromoacetyl
Reactive toward: amino and sulfhydryl groups
• NHS ester reacts with primary amines at pH 7-9 to form a stable amide bond
• Bromacetyl group reacts with sulfhydryl groups at pH > 7.5 to form stable thioether bonds
• Non-cleavable
• Water-insoluble (dissolve first in DMF or DMSO)
• Spacer maintains peptide-like character in the crosslinked species
• Resulting crosslink is susceptible to acid hydrolysis
• Useful for preparing cyclic peptides and peptide conjugates

Product References:

Crosslinker Application Guide -- search for recent literature references for this product

DTSSP (3,3'-dithiobis(sulfosuccinimidyl propionate)) Thermo Scientific™

Thermo Scientific Pierce DTSSP is a popular water-soluble crosslinker that contains amine-reactive NHS-ester ends around an 8-atom spacer arm, whose central disulfide bond can be cleaved with reducing agents.

DTSSP is 3,3'-Dithiobis(sulfosuccinimidylpropionate), which contains an amine-reactive N-hydroxysulfosuccinimide (sulfo-NHS) ester at each end of an 8-carbon spacer arm. Sulfo-NHS esters react with primary amines at pH 7-9 to form stable amide bonds, along with release of the N-hydroxysulfosuccinimide leaving group. Proteins, including antibodies, generally have several primary amines in the side chain of lysine (K) residues and the N-terminus of each polypeptide that are available as targets for sulfo-NHS-ester crosslinking reagents. DSS, the non-sulfonated analog of DTSSP is also available for applications that require a membrane-permeable crosslinker.

Features of DTSSP:

Reactive groups: sulfo-NHS ester (both ends)
Reactive towards: amino groups (primary amines)
• Sulfo-NHS ester reacts rapidly with any primary amine-containing molecule
• Disulfide bond in the spacer arm is readily cleaved by 10-50 mM DTT or TCEP at pH 8.5
• Spacer arm also easily cleaved with reducing SDS-PAGE sample loading
• Cleavable crosslinker allows separation of crosslinked products
• Water-soluble; compare with DSP
• Membrane-impermeable, allowing for cell surface labeling

Applications:
• Chemical crosslinking of cell surface proteins prior to cell lysis and immunoprecipitation
• 'Fix' protein interactions to allow identification of weak or transient protein interactions
• Protein crosslinking to create bioconjugates via single-step reactions
• Immobilize proteins onto amine-coated surfaces

Properties of DTSSP
• Alternative names: Sulfo-DSP
• Molecular formula: C14H14O14N2S4Na2
• Molecular weight: 608.51
• Spacer arm length: 12.0 Å (8 atoms)
• Reactive groups: Sulfo-NHS esters, react with primary amines at pH 7.0–9.0

DTSSP Specifications
We manufacture DTSSP to the highest specifications to produce the most specific bioconjugates, ensure the integrity of your data and to provide you with the highest degree of consistency. Each lot of DTSSP is tested to meet the following minimum specifications:
Solubility: > 6 mg/mL in DI water
Identity: IR scan shows only peaks characteristic of the structure and functional groups of DTSSP
Purity: > 80% by quantitative NMR (the highest standard for crosslinker purity)

Product References:
Crosslinker Application Guide -- search for recent literature references for this product

Related Products
DTSSP (3,3'-dithiobis(sulfosuccinimidyl propionate))
DSP (dithiobis(succinimidyl propionate)), Lomant's Reagent
DSS (disuccinimidyl suberate)

SM(PEG)12 (PEGylated, long-chain SMCC crosslinker) Thermo Scientific™

Thermo Scientific Pierce SM(PEG)n is a series of amine-to-sulfhydryl crosslinkers that differ in length from 17.6 to 95.2 angstroms as a result of polyethylene glycol spacer arms containing n equals 2 to 24 ethylene glycol units.

Features of SM(PEG)12:

Reactive groups: NHS ester and maleimide
Reactive toward: amino and sulfhydryl groups
• Irreversibly crosslink proteins or peptides by flexible PEG spacer arms
• Polyethylene glycol spacer arms help maintain conjugate solubility
• Pure compound with defined structure and molecular weight, ensuring reproducible protein-modification effects
• PEG spacer provides unique advantages, including increased stability, reduced tendency toward aggregation and reduced immunogenicity
• Ideal for performing controlled, two-step protein conjugations (see Sulfo-SMCC)

SM(PEG)n NHS- and maleimide-activated PEG compounds for crosslinking between primary amines (NH2) and sulfhydryl (SH) groups in proteins and other molecules. A complete set of compounds are offered, each having the same heterobifunctional structure (NHS-PEGn-Maleimide) but differing in the number of discrete ethylene glycol units ( n = 2, 4, 6, 8, 12 or 24). The N-hydroxysuccinimide ester (NHS) group reacts specifically and efficiently with lysine and N-terminal amino groups at pH 7-9 to form stable amide bonds. The maleimide group reacts with reduced sulfhydryls at pH 6.5-7.5 to form stable thioether bonds.

Alternative Names for SM(PEG)n:
• NHS-dPEG-Mal
• Mal-PEG NHS ester
• NHS-PEO-Maleimide
• Succinimidyl-dPEG-maleimide
• Succinimide-Maleimide PEG

Why use crosslinkers with polyethylene glycol (PEG) spacer arms?
PEG-containing reagents have been used to modify proteins to provide specific advantages. Protein PEGylation can improve the stability of the modified protein, protect it from proteolytic digestion, increase its half life in a biological application, mask it from causing an immunogenic response, decrease its antigenicity or potential toxicity, improve its solubility, diminish the potential for aggregation, and minimize interference for both in vitro and in vivo applications. Polyethylene glycol has these effects because it is nontoxic, nonimmunogenic, hydrophilic, water soluble and highly flexible.

Advantages of discrete-length polyethylene glycol compounds:
These reagents are specially synthesized, resulting in homogeneous compounds of defined molecular weight, characterized by discrete chain lengths, providing a greater ability to optimize and characterize surface protein modifications. Typical preparations of PEG compounds are a heterogeneous mixtures composed of a distribution of chain lengths with a specified average molecular weight or approximate number of PEG subunits.

Related Products
SM(PEG)6 (PEGylated, long-chain SMCC crosslinker)
SM(PEG)8 (PEGylated, long-chain SMCC crosslinker)
SM(PEG)24 (PEGylated, long-chain SMCC crosslinker)

SMPT (4-succinimidyloxycarbonyl-alpha-methyl-α(2-pyridyldithio)toluene) Thermo Scientific™

Thermo Scientific Pierce SMPT is a long-chain crosslinker for amine-to-sulfhydryl conjugation via NHS-ester and pyridyldithiol reactive groups that form strong (hindered) yet cleavable (reducible) disulfide bonds with cysteine sulfhydryls.

Features of SMPT:

Reactive groups: NHS ester and pyridyldithiol
Reactive towards: amino and sulfhydryl groups
• Cleavable with disulfide reducing agents
• Water-insoluble (dissolve first in DMF or DMSO)
• Contains a hindered disulfide bond; has formed immunotoxins with improved stability
• In vitro, an SMPT conjugate was as effective as conjugates formed with SPDP and 2-IT
• Compare to other varieties of SPDP-type reagents, including pegylated forms

Product References:
Crosslinker Application Guide -- search for recent literature references for this product

Related Products
SPDP (succinimidyl 3-(2-pyridyldithio)propionate)

DSS (disuccinimidyl suberate), No-Weigh™ Format Thermo Scientific™

Thermo Scientific Pierce DSS, also called disuccinimidyl suberate, is noncleavable and membrane permeable crosslinker that contains an amine-reactive N-hydroxysuccinimide (NHS) ester at each end of an 8-carbon spacer arm.

NHS esters react with primary amines at pH 7-9 to form stable amide bonds, along with release of the N-hydroxysuccinimide leaving group. Proteins, including antibodies, generally have several primary amines in the side chain of lysine (K) residues and the N-terminus of each polypeptide that are available as targets for NHS-ester crosslinking reagents. DSS is first dissolved in an organic solvent such as DMF or DMSO, then added to the aqueous crosslinking reaction. BS3, the water soluble analog of DSS is also available for applications that require a hydrophilic crosslinker (e.g. to effect cell-surface crosslinking). DSS and BS3 have essentially identical crosslinking activity toward primary amines.

Features of disuccinimidyl suberate:

Reactive groups: NHS ester (both ends)
Reactive towards: amino groups (primary amines)
• Amine reactive sulfo-NHS ester reacts rapidly with any primary amine-containing molecule
• Membrane-permeable, allowing for intracellular crosslinking
• High purity, crystalline reagent can be used to create high-purity conjugates
• Non-cleavable
• Water-insoluble (dissolve first in DMF or DMSO); compare to BS3 (Sulfo-DSS)

Applications:
• Chemical crosslinking of intracellular proteins prior to cell lysis and immunoprecipitation
• 'Fix' protein interactions to allow identification of weak or transient protein interactions
• Protein crosslinking to create bioconjugates via single-step reactions
• Immobilize proteins onto amine-coated surfaces

Properties of DSS
• Alternative names: Disuccinimidyl suberate
• Molecular formula: C16H20N2O8
• Molecular weight: 368.35
• Spacer arm length: Å (8 atoms)
• CAS number: 68528-80-3
• Reactive groups: NHS esters, react with primary amines at pH 7.0–9.0

DSS Specifications
We manufacture DSS to the highest specifications to produce the most specific bioconjugates, ensure the integrity of your data and to provide you with the highest degree of consistency. Each lot of DSS crosslinker is tested to meet the following minimum specifications:
Identity: IR scan shows only peaks characteristic of the structure and functional groups of DSS
Purity: > 90% by quantitative NMR (the highest standard in crosslinking purity)
Solubility: > 9.2 mg/mL in DMF and DMSO, clear and colorless solution

Product References
Crosslinker Application Guide -- search for recent literature references for this product

Related Products
BS3 (bis(sulfosuccinimidyl)suberate)

PMPI (p-maleimidophenyl isocyanate) Thermo Scientific™

Thermo Scientific Pierce PMPI is a maleimide-and-isocyanate crosslinker for attaching compounds to sulfhydryl groups (cysteines) after conjugating the linker to hydroxyl groups on a molecule by urethane (carbamate) bonds.

Features of PMPI:

Reactive groups: maleimide and isocyanate
Reactive towards: sulfhydryl and hydroxyl groups
• Short (8.7A), sulfhydryl-to-hydroxyl crosslinker with an aromatic spacer arm (noncleavable)
• Maleimide group reacts with sulfhydryl groups to form stable thioether linkages
• Isocyanate reacts with non-aqueous hydroxyl groups to form carbamate linkage
• Excellent tool for conjugating hydroxyl-containing compounds such as steroids and vitamins
• Targets and reaction solutions must not contain primary amines

Pierce™ TCEP-HCl, No-Weigh™ Format Thermo Scientific™

Thermo Scientific Pierce TCEP-HCl is a potent, versatile, odorless, thiol-free reducing agent with broad application to protein and other research involving reduction of disulfide bonds. The unique compound is easily soluble and very stable in many aqueous solutions. TCEP reduces disulfide bonds as effectively as dithiothreitol (DTT), but unlike DTT and other thiol-containing reducing agents, TCEP does not have to be removed before certain sulfhydryl-reactive crosslinking reactions.

Thermo Scientific No-Weigh products are specialty reagents provided in a pre-aliquoted format. The pre-weighed packaging prevents the loss of reagent reactivity and contamination over time by eliminating the repetitive opening and closing of the vial. The format enables use of a fresh vial of reagent each time, eliminating the hassle of weighing small amounts of reagents and reducing concerns over reagent stability.

Features of TCEP-HCl:

Odorless—Unlike DTT or BME, TCEP is odor-free, so it can reduce proteins conveniently on the bench; contributes to a healthier lab environment
Specific—selective and complete reduction of even the most stable water-soluble disulfides over a wide pH range
Simple—effective reduction at room temperature and pH 5 in less than five minutes
Stable—it's inherent stability and resistant to air oxidation eliminates the need for any special precautions while handling or storing; non-volatile and non-reactive toward other functional groups found in proteins
Efficient—For most applications, 5 to 50 mM TCEP provides sufficient molar excess to effectively reduce peptide or protein disulfide bonds within a few minutes at room temperature.
Compatible—With TCEP, removal of the reducing agent is not necessary prior to most applications, (e.g. histidine-tagged protein purification, maleimide conjugations).

Considerations for use of TCEP:
• TCEP is generally very soluble in aqueous buffers at nearly any pH. Therefore, working concentrations and 10X stock solutions may be readily prepared in most aqueous buffers.
• TCEP is stable in aqueous, acidic, and basic solutions. When TCEP is dissolved directly in water, the resulting pH is approximately 2.5.
• TCEP is not particularly stable in phosphate buffers, especially at neutral or alkaline pH. Therefore, if TCEP is to be used in PBS buffers, prepare the working solution immediately before use.
• TCEP may be used as a substitute for DTT or 2-mercaptoethanol (2-ME) in sample loading buffer for SDS-PAGE; use a final concentration of 50 mM TCEP.
• Because TCEP is charged in solution, it is not compatible for use in isoelectric focusing.

EGS (ethylene glycol bis(succinimidyl succinate)) Thermo Scientific™

Thermo Scientific Pierce EGS is a crosslinker that contains amine-reactive NHS-ester ends around a 12-atom spacer arm, which can be cleaved by treatment with hydroxylamine at pH 8.5.

Features of the EGS crosslinker:

Reactive groups: NHS ester (both ends)
Reactive towards: amino groups (primary amines)
• Water-insoluble (dissolve first in DMF or DMSO); compare to Sulfo-EGS
• Membrane-permeable, allowing for intracellular crosslinking
• Crosslinks formed are reversible at pH 8.5 using hydroxylamine for 3 to 6 hours at 37°C
• Lactose dehydrogenase retained 60% of its activity after reversible crosslinking with EGS

Product References:
Crosslinker Application Guide -- search for recent literature references for this product

Related Products
EGS (ethylene glycol bis(succinimidyl succinate))

DTME (dithiobismaleimidoethane) Thermo Scientific™

Thermo Scientific Pierce DTME is a mid-length, maleimide crosslinker for reversible covalent conjugation between sulfhydryl groups (e.g., protein or peptide cysteines) by reduction of the disulfide bond in the center of the spacer arm.

Features of DTME:

Reactive groups: maleimide (both ends)
Reactive towards: sulfhydryl groups
• Long (13.3A), cleavable, sulfhydryl-to-sulfhydryl crosslinker, composed of maleimide groups and 11-atom disulfide spacer arm
• Water-insoluble—dissolve first in DMF or DMSO, then add to aqueous reaction buffers
• Cleavable by reduction of disulfide spacer arm with DTT, TCEP or other reducing agent

Product References:
Crosslinker Application Guide -- search for recent literature references for this product

Related Products
BMH (bismaleimidohexane)

AMAS (N-α-maleimidoacet-oxysuccinimide ester) Thermo Scientific™

Thermo Scientific Pierce AMAS is an amine-to-sulfhydryl crosslinker that contains NHS-ester and maleimide reactive groups at opposite ends of a very short spacer arm (4.4 angstroms).

Features of AMAS:

Reactive groups: NHS ester and maleimide
Reactive towards: amino and sulfhydryl groups
• Shortest of aliphatic spacer series which includes AMAS, BMPS, GMBS and EMCS
• NHS ester end couples with primary amines at pH 7-9, forming stable amide bonds
• Maleimide reacts with -SH groups at pH 6.5-7.5, forming stable thioether linkages
• Non-cleavable
• Water-insoluble (dissolve first in DMF or DMSO)
• Very short aliphatic spacer has low potential for eliciting an immune response
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