NHS-dPEG®₄-biotin

NHS-dPEG®4-biotin, product number 10200, is a constant favorite among our customers. It has been used or featured in numerous publications over the years since it was first introduced. Biotin is normally poorly soluble in water, but the amphiphilic dPEG® linker imparts excellent solubility in water or aqueous buffer and in organic solvent. The N-hydroxysuccinimidyl (NHS) moiety of NHS-dPEG®4-biotin reacts with free primary amines, such as the amines on the side chain of lysine, forming a stable amide bond. The biotin moiety is used in labeling experiments, supramolecular construction, affinity chromatography, the creation of biotinylated antibodies, and many other applications.

Biotinylation with NHS-dPEG®4-biotin
In times past, aminocaproic acid was coupled to biotin and activated with NHS to form the widely used NHS-LC-biotin, but NHS-LC-biotin is highly problematic, as it is insoluble in water and must be dissolved in organic media prior to use. When coupled to proteins, the linker's high degree of hydrophobicity can cause rapid aggregation of the proteins. A variant of NHS-LC-biotin, known as Sulfo-NHS-LC-biotin, is water soluble, but it still causes rapid aggregation of proteins.

NHS-dPEG®4-biotin does not cause protein aggregation. This permits experiments that require a high degree of biotin labeling without concerns about loss of protein due to aggregation. Dissolution of the product in organic solvent is not required for conjugation, thus providing easier reaction setup and greater reaction reproducibility. The dPEG® linker in PN10200 is slightly longer than the aminocaproic acid linker in LC-biotin; therefore, dPEG®4-biotin provides better access of the biotin to the avidin or streptavidin binding pocket compared to LC-biotin.

PN10200 is part of a family of dPEG®-biotin reagents that provide different lengths. The active esters can be either NHS or 2,3,5,6-tetrafluorophenol (TFP), which is more reactive towards primary amines and more stable in water or aqueous buffer. The optimum pH for reaction with NHS esters is in the range of 7.2 – 7.5, while for TFP esters, the optimum reaction pH range is slightly higher.

Application References:

Hermanson, G. T. Chapter 11, (Strept)avidin-Biotin Systems. Bioconjugate Techniques, 3rd edition. Academic Press: New York, 2013, 465-505.
Hermanson, G. T. Chapter 18, PEGylation and Synthetic Polymer Modification. Bioconjugate Techniques, 3rd edition. Academic Press: New York, 2013, 787-838.

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