Thermo Scientific Pierce Sulfo-NHS (N-hydroxysulfosuccinimide) and its uncharged analog NHS (N-hydroxysuccinimide) are chemical modification reagents for converting carboxyl groups to amine-reactive Sulfo-NHS esters for application in a variety of bioconjugation and crosslinking methods.
Derivatives are easily synthesized by mixing the Sulfo-NHS with a carboxyl-containing molecule and a dehydrating agent such as the carbodiimide EDC (EDAC). The method is the basis for generating many types of protein labeling reagents, including amine-reactive fluorescent dyes, biotin affinity tags and pegylation compounds.
- Efficiency of EDC-mediated coupling is increased in the presence of Sulfo-NHS
- Amine-reactive Sulfo-NHS esters can be created on any carboxyl-containing molecule
- Sulfo-NHS derivatives are water-soluble, so crosslinking can be done in physiologic solutions
- High-purity, crystalline Sulfo-NHS can be used to create high-purity activated derivatives
- Improve efficiency of EDC coupling reactions
- Convert carboxyls to amine-reactive Sulfo-NHS esters
- Crosslink proteins to carboxyl-coated beads or surfaces more efficiently
- Activate nanoparticles with amine-reactive Sulfo-NHS esters
Activation and crosslinking with EDC and Sulfo-NHS. EDC by itself is not particularly efficient in crosslinking because failure to react quickly with an amine will result in hydrolysis and regeneration of the carboxyl.EDC reacts with a carboxyl group on molecule #1, forming an amine-reactive O-acylisourea intermediate. This intermediate may react with an amine on molecule #2, yielding a conjugate of the two molecules joined by a stable amide bond. However, the intermediate is also susceptible to hydrolysis, making it unstable and short-lived in aqueous solution. The addition of Sulfo-NHS (5 mM) stabilizes the amine-reactive intermediate by converting it to an amine-reactive Sulfo-NHS ester, thus increasing the efficiency of EDC-mediated coupling reactions (Ref.1,2). The amine-reactive Sulfo-NHS ester intermediate has sufficient stability to permit two-step crosslinking procedures, which allows the carboxyl groups on protein #2 to remain unaltered.
Properties of NHS and Sulfo-NHS
||Store at 4°C, protect from moisture, use only fresh solutions
Specifications for NHS and Sulfo-NHS:
We manufacture N-hydroxysulfosuccinimide to the highest possible specifications to produce the most specific bioconjugates, to ensure the integrity of your data and to provide you with the highest degree of consistency. Each lot of Sulfo-NHS is tested to meet the following minimum specifications:
- Purity – Greater than 95% by quantitative NMR (the highest standard for crosslinker purity);
Average lot purity is greater than 99%! See the Sulfo-NHS NMR data.
- Solubility – Sample dissolves at 2 mg/ml in deionized water to yield a clear, colorless solution
- Identity – IR scan shows only peaks characteristic of N-hydroxysulfosuccinimide
- Zhao C. et al. (2010) Effects of a lubricin-containing compound on the results of flexor tendon repair in a canine model in vivo. J. Bone Joint Surg. Am. 92, 1453-61.
- Anderson G. L. et al. (2010) Assessing lead time of selected ovarian cancer biomarkers: A nested case-control study. J Natl Cancer Inst. 102, 26-38.
- Ni J. et al. (2009) Plasma protein pentosidine and carboxymethyllysine, biomarkers for age-related macular degeneration. Mol. Cell. Proteomics. 8, 1921-33.
- Jaetao J. E. et al. (2009) Enhanced leukemia cell detection using a novel magnetic needle and nanoparticles. Cancer Res. 69, 8310-6.
- Marintcheva B. et al. (2008) Acidic c-terminal tail of the ssdna-binding protein of bacteriophage t7 and ssdna compete for the same binding surface. PNAS. 105, 1855-60.
Crosslinker Selection Guide
NHS and Sulfo-NHS FAQ
Protocol for efficient two-step coupling of proteins using EDC and Sulfo-NHS
Tech Tip #3: Determine the activity of NHS esters
Tech Tip #46: Preferentially label the amino-terminus of peptides
Sulfo-NHS is available in bulk quantities for manufacturing applications.