NHS-dPEG®₁₂-biotin

Product number 10198, NHS-dPEG®12-biotin, is a longer version of the constantly popular product number 10200, NHS-dPEG®4-biotin. This product has been used and featured in numerous publications 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®12-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, pull-down assays, the creation of biotinylated antibodies, and many other applications.

NHS-LC-biotin (biotin coupled to aminocaproic acid and activated with N-hydroxysuccinimide) and NHS-LC-LC-biotin (with two aminocaproic acid chains) are widely used biotinylation reagents. However, these two reagents are problematic for biochemical applications. Insoluble in water, both reagents must be dissolved in organic solvent immediately prior to use. When coupled to proteins, the linkers' high degree of hydrophobicity causes rapid aggregation and deactivation 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®12-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 PN10198 is longer than the linkers in LC-biotin and LC-LC-biotin. Consequently, dPEG®12-biotin provides better access of the biotin to the avidin or streptavidin binding pocket compared to these older, hydrophobic reagents.

PN10198 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). Compared to NHS ester, TFP esters are 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.

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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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