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EZ-Link Alkoxyamine-PEG4-SS-PEG4-Biotin


Glycoprotein biotinylation via a cleavable, pegylated spacer arm.

EZ-Link Alkoxyamine-PEG4-SS-PEG4-Biotin

Thermo Scientific EZ-Link Alkoxyamine-PEG4-SS-PEG4-Biotin is a reagent containing a long polyethylene glycol (PEG) spacer and cleavable disulfide bond for reversibly biotinylating at carbohydrate groups that have been oxidized to aldehydes.

Aminooxy-biotin reagents such as this one are useful for biotinylating glycoproteins and other molecules that have oxidizable polysaccharides groups. The alkoxyamine group (also called an aminooxy or aminoxy group) conjugates to aldehydes of oxidized sugars. EZ-Link Alkoxyamine-PEG-Biotin reagents contain a multi-functional extended spacer arm that is a flexible, non-immunogenic hydrophilic polyethylene glycol (PEG), which imparts water solubility to labeled molecules. Consequently, antibodies labeled with pegylated biotin reagents exhibit less aggregation when stored in solution compared to antibodies labeled with reagents having only hydrocarbon spacers. Alkoxyamine-PEG4-SS-PEG4-Biotin has a disulfide bond in the spacer and two PEG chains. The disulfide bond enables cleavage of the biotin group from the labeled proteins, which is especially useful when affinity purifying biotinylated proteins using immobilized avidin, streptavidin or Thermo Scientific NeutrAvidin Protein.

Highlights:

  • Glycoprotein labeling – biotinylate glycosylated proteins at sialic acid residues for detection or purification using streptavidin probes or resins
  • Cell surface labeling – biotinylate and isolate cell surface glycoproteins; reagent does not permeate membranes of whole cells
  • Aldehyde-reactive – reacts with aldehydes formed by periodate-oxidation of sugar groups
  • Alkoxyamine-activated – aminooxy group forms more stable linkages than hydrazide reagents
  • Pegylated – spacer arm contains a hydrophilic, 2 x 4-unit, polyethylene glycol (PEG) group
  • Enhances solubility – pegylation imparts water solubility to the biotinylated molecule, helping to prevent aggregation of biotinylated antibodies stored in solution
  • Solubility – usually dissolved in DMSO to make concentrated stock solution
  • Very long reach – spacer arm (total length added to target) is 56.6 angstroms
  • Cleavable – disulfide bond in spacer arm allows the biotin label to be removed using reducing agents such as DTT; a pegylated sulfhydryl group remains attached to the molecule

Product Details:

Chemical structure of Alkoxyamine-PEG4-SS-PEG4-Biotin
Chemical structure of Alkoxyamine-PEG4-SS-PEG4-Biotin. This aminooxy-activated biotinylation reagent labels glycoproteins and other molecules having carbohydrates that can be oxidized with periodate to expose aldehydes. For more information, see our review of Alkoxyamine Reaction Chemistry.

Properties of Alkoxyamine-PEG4-SS-PEG4-Biotin. This aldehyde-reactive biotinylation reagent has a long spacer arm that contains two polyethylene glycol segments and a cleavable disulfide bond.
Alternative names Biotinamido-PEG4-SS-PEG4-hydroxylamine; Aminooxy-PEG4-SS-PEG4-Biotin
SMILES structure O=C(CCCCC1SCC(N2)C1NC2=O)NCCOCCOCCOCCOCCC(NCCSSCCC(NCCOCCOCCOCCON)=O)=O
Molecular formula C34H66N4O14S
Molecular weight 845.10
Spacer arm length 56.6 angstroms
Mass added to target 827.08 daltons
Form Viscous semi-solid
Solubility Dissolve in DMSO to make 250mM stock
Storage conditions -20°C protected from moisture
Reactive groups Alkoxyamine (Aminooxy), reacts with aldehydes at near-neutral pH
Cleave with 50mM DTT or other reducing agents

We manufacture biotin reagents to ensure the highest possible overall product integrity, consistency and performance for the intended research applications.

Biotinylation reagents differ in reactivity, length, solubility, cell permeability and cleavability. Hydrazides and alkoxyamines are two types of carbonyl-reactive groups. Alkoxyamines (–O-NH2) react specifically with aldehyde groups in near-neutral conditions to form stable oxime linkages. The reaction is more efficient in the presence of aniline (Part No. 88944). Alternatively, alkoxyamines can be conjugated to carboxylic acids using EDC carbodiimide chemistry.

Reactive aldehyde groups can be generated in glycoproteins and other polysaccharide compounds by oxidation of constituent sugar diols using sodium periodiate (Part No. 20504). Sialic acid residues are common components of protein glycosylation and are easily converted to aldehydes with 1mM NaIO4.

General References:

  1. Dirksen, A., et al. (2006). Nucleophilic catalysis of oxime ligation. Angew Chem Int Ed 45:7581-4.
  2. Dirksen, A. and Dawson, P. (2008). Rapid oxime and hydrazone ligations with aromatic aldehydes for biomolecular labeling. Bioconjugate Chem 19:2543-8.
  3. Lempens, E., et al. (2009). Efficient and chemoselective surface immobilization of proteins by using aniline-catalyzed oxime chemistry. Chembiochem 10:658-62.
  4. Mahal, L.K., et al. (1997). Engineering chemical reactivity on cell surfaces through oligosaccharide biosynthesis. Science 276:1125-8.
  5. Zeng, Y., et al. (2009). High efficiency labeling of glycoproteins on living cells. Nat Methods 6:207-9.

Related Resources:

Biotinylation Reagents Selection Guide
Review of Biotinylation Methods and Applications
Review of Glycosylation and Glycoprotein Methods
Chemistry of Crosslinking (and Labeling Reagents)
Review of PEGylation

Related Products:

Alkoxyamine-PEG12-Biotin
Sodium meta-Periodate
All Biotinylation Reagents
Avidin, Streptavidin, NeutrAvidin Affinity Resins
PEGylation Reagents


Ordering Information
 
Product # Description Pkg. Size Instructions MSDS CofA Price
26138 EZ-Link Alkoxyamine-PEG4-SS-PEG4-Biotin
Formulation: Alkoxyamine-activated biotin with a poly(ethyleneglycol) and dithiol spacer
Sufficient For: Approx. 12 uses to label a total of 12mg of IgG (oxidized glycoprotein) using typical conditions
50mg Product Instructions for product #26138 EZ-Link Alkoxyamine-PEG4-SS-PEG4-Biotin MSDS for product #26138 EZ-Link Alkoxyamine-PEG4-SS-PEG4-Biotin Certificate of Analysis for product #26138 EZ-Link Alkoxyamine-PEG4-SS-PEG4-Biotin $297.00

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Instructions | MSDS | CofA
Product Instructions | MSDS | CofA  

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