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EZ-Link NHS-SS-PEG4-Biotin

A long-arm, pegylated biotinylation reagent with a cleavable disulfide bond.

EZ-Link NHS-SS-PEG4-Biotin

Thermo Scientific EZ-Link NHS-SS-PEG4-Biotin includes disulfide and polyethylene glycol (PEG) groups in its spacer arm to provide biotinylation that is cleavable and which enhances solubility.

NHS-SS-PEG4-Biotin enables simple and efficient biotin labeling of antibodies, proteins and other primary amine-containing macromolecules. The N-hydroxysuccinimide ester (NHS ester) group reacts specifically and efficiently with protein and peptide lysine (K) and N-terminal amino groups at pH 7-9 to form stable amide bonds. The hydrophilic polyethylene glycol (PEG) spacer arm imparts water solubility that is transferred to the biotinylated molecule, thus reducing aggregation of labeled proteins stored in solution. The integral disulfide bond enables the biotin group to be cleaved from the labeled molecule using DTT or other reducing agents, providing for efficient recovery of avidin-bound protein complexes in affinity-based assays.


  • Protein labeling – biotinylate antibodies or other proteins for detection or purification using streptavidin probes or resins
  • Amine-reactive – reacts with primary amines (-NH2), such as the side-chain of lysines (K) or the amino-termini of polypeptides
  • Cleavable – disulfide bond in spacer arm allows the biotin label to be removed using reducing agents such as DTT; only a small sulfhydryl group remains attached to the molecule
  • Pegylated – spacer arm contains a hydrophilic, 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
  • Long reach – spacer arm (total length added to target) is 37.9 angstroms; this reduces steric hindrance when binding to avidin molecules

Product Details:

Chemical structure of NHS-SS-PEG4-Biotin
Chemical structure of NHS-SS-PEG4-Biotin. This biotinylation reagent labels antibodies and other proteins and molecules that contain primary amines (e.g., side-chain of lysine). For more information, see our review of NHS Ester Reaction Chemistry.

Properties of EZ-Link NHS-SS-PEG4-Biotin. This amine-reactive compound has a cleavable disulfide bond, a hydrophilic polyethylene glycol (PEG) segment and is cell membrane permeable.
Alternative names NHS-SS-PEO4-Biotin
Molecular formula C30H49N5O11S3
Molecular weight 751.94
Spacer arm length 37.9 angstroms
Mass added to target 636.85 daltons
Form White to off-white solid
Storage conditions -20°C with desiccant
Reactive groups N-hydroxysuccinimide ester, reacts with primary amines at pH 7.0-9.0
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.

N-Hydroxysulfosuccinimide (NHS) esters of biotin are the most popular type of biotinylation reagent. NHS-activated biotins react efficiently with primary amino groups (-NH2) in alkaline buffers to form stable amide bonds. Proteins (e.g., antibodies) typically have several primary amines that are available as targets for labeling, including the side chain of lysine (K) residues and the N-terminus of each polypeptide.

Varieties of biotin NHS-ester reagents differ in length, solubility, cell permeability and cleavability. Non-sulfonated NHS-biotins are cell permeable but must be dissolved in organic solvent such as DMSO or DMF. Sulfo-NHS biotins (and those with pegylated spacers) are directly water soluble but not membrane permeable. Varieties containing disulfide bonds can be cleaved using reducing agents, enabling the biotin group to be disconnected from the labeled protein.


  1. Causeret, M., et al. (2005).  Mol. Biol. Cell. 16, 2168-2180.
  2. Morimoto, S., et al. (2005).  J. Biol. Chem. 280, 2220-2228.
  3. Trotti, D., et al. (2001).  J. Biol. Chem. 276, 576-582.
  4. Daniels, G.M. and Amara, S.G. (1998).  Methods. Enzymol. 296, 307-318.
  5. Soukup, G.A., et al. (1995).  Bioconjugate Chemistry. 6, 135-138.
  6. Shimkus, M., Levy, J. and Herman, T. (1985).  Proc. Natl. Acad. Sci. 82, 2593-2597.
  7. Duhamel, R.C. and Johnson, D.A. (1985).  J. Histochem. Cytochem. 33, 711-714.

Related Resources:

Biotinylation Reagents Selection Guide
Review of Biotinylation Methods and Applications
Review of Pegylation (Polyethylene Glycol)
Chemistry of Crosslinking (and Labeling Reagents)

Related Products:

Pierce Cell Surface Protein Isolation Kit
All Biotinylation Reagents
Avidin, Streptavidin, NeutrAvidin Affinity Resins

Ordering Information
Product # Description Pkg. Size Instructions MSDS CofA Price
21442 EZ-Link NHS-SS-PEG4-Biotin
Formulation: NHS ester of biotin with an ethyl-1,3-dithioproprionate and tetraethyleneglycol spacer
Sufficient For: Approx. 25 uses to label a total of 443mg of IgG using typical conditions
50mg Product Instructions for product #21442 EZ-Link NHS-SS-PEG4-Biotin MSDS for product #21442 EZ-Link NHS-SS-PEG4-Biotin Certificate of Analysis for product #21442 EZ-Link NHS-SS-PEG4-Biotin $320.00

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Instructions | MSDS | CofA
Product Instructions | MSDS | CofA  

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