Chemoselective ligation refers to the use of mutually specific pairs of conjugation reagents. Staudinger ligation reagents are pairs of metabolic or chemical labeling compounds that have azide and phosphine groups, respectively. These reactive groups recognize each other but not any natural or endogenous biomolecules in typical samples. When respective pairs of target biomolecules are labeled with these two groups, they will conjugate to one another with high specificity when combined. This specialized form of crosslinking is most often used for in vivo metabolic labeling.

NHS-Azide Staudinger Ligation Reagents - These NHS-ester compounds react to form covalent bonds with primary amines (e.g., side chain of lysine residues or aminosilane-coated surfaces). The azide (N3) group reacts with phosphine-labeled molecules by a mechanism known as Staudinger chemistry, enabling efficient and specific conjugation of derivatized molecules in biological samples. In combination with different phosphine-activated compounds, these NHS-Azide reagents facilitate a variety of labeling and crosslinking experimental strategies for investigating protein interactions and cellular pathways.

Properties of NHS-Azide reagents

These compounds are identical except for the their overall length as a result of different numbers of polyethylene glycol groups (PEG units).

  NHS-Azide NHS-PEG4-Azide
Chemical formula C6H6N4O4 C15H24N4O8
Molecular weight 198.14 388.37
Spacer arm length 2.5 angstroms 18.9 angstroms
Form white to off-white powder colorless to pale yellow liquid
SMILES structures O=C(ON1C(CCC1=O)=O)CN=[N+]=[N-]
Storage conditions -20°C with desiccant, protect from moisture and light, use only fresh solutions
Reactive groups N-hydroxysuccinimide ester, reacts with primary amines at pH 7.0-9.0;
Azide, reacts with phosphine groups

Products for crosslinking using Staudinger chemistry