Thermo Scientific EZ-Link NHS-PEG4-Biotin is a pegylated, water-soluble reagent for simple and efficient biotin labeling of antibodies, proteins and other primary amine-containing macromolecules.
NHS-PEG4-Biotin is a long (29.0A), pegylated, water-soluble, NHS-ester biotinylation reagent to label amines and maximize solubility of antibodies and other proteins. The N-hydroxysuccinimide ester (NHS) group reacts specifically and efficiently with lysine and N-terminal amino groups 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 PEG spacer arm also gives the reagent a long and flexible connection to minimize steric hindrance for binding to avidin molecules.
- 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
- 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
- Irreversible – forms permanent amide bonds; spacer arm cannot be cleaved
- Long reach – spacer arm (total length added to target) is 29 angstroms; this reduces steric hindrance when binding to avidin molecules
|Chemical structure of NHS-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-PEG-Biotin Reagents. These compounds have polyethylene glycol (PEG) spacer arms that enhance the reach and solubility of labeled molecules.
|Spacer arm length
|Mass added to target
||White to off-white solid
||White to off-white semi-solid
||10mg/mL in water
||Best to make DMF or DMSO stock
-20°C with desiccant
-20°C with desiccant
|SMILES structure (PEG4)
||N-hydroxysuccinimide ester, reacts with primary amines at pH 7.0-9.0
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.
- Altin, J.G., et al. (1995. A one-step procedure for biotinylation and chemical crosslinking of lymphocyte surface and intracellular membrane-associated molecules. Anal. Biochem. 224, 382-9.
- Gretch, D.R., et al. (1987) The use of biotinylated monoclonal antibodies and streptavidin affinity chromatography to isolate herpes virus hydrophobic proteins or glycoproteins. Anal. Biochem. 163, 270-7.
- Manning, J., et al. (1977) A method for gene enrichment based on the avidin-biotin interaction. Application to the Drosophila ribosomal RNA genes. Biochemistry. 16, 1364-70.
- Updyke, T.V. and Nicolson, G.L. (1984) Immunoaffinity isolation of membrane antigens with biotinylated monoclonal antibodies and immobilized streptavidin matrices. J. Immunol. Meth. 73, 83-95.
- Luo, Y., et al. (2003) Mol. Cell. Biol. 23, 2600-7.
- Nielsen, P.A., et al. (2002) J. Biol. Chem. 277, 38272-83.
- Xu, G. et al. (2004) J. Cell Sci. 117, 3207-3219.
- Vaidya, V.S., et al. (2006) Am. J. Physiol. in press.
- Nikitina, T. and Woodcock, C.L. (2004) J. Cell Biol. 166, 161-165.
Biotinylation Reagents Selection Guide
Review of Biotinylation Methods and Applications
Review of Pegylation (Polyethylene Glycol)
Chemistry of Crosslinking (and Labeling Reagents)
NHS-PEG12-Biotin – very long (56.0 angstrom) spacer arm variant
Sulfo-NHS-Biotin and Kits – 13.5 angstrom spacer arm
Sulfo-NHS-LC-Biotin and Kits – 22.4 angstrom spacer arm
Sulfo-NHS-SS-Biotin and Kits – 13.5 angstrom spacer arm, cleavable
All Amine-Reactive Biotinylation Reagents
All Biotinylation Reagents
Biotin Quantitation Reagents and Kits
Avidin, Streptavidin, NeutrAvidin Affinity Resins