Thermo Scientific EZ-Link Sulfo-NHS-LC-Desthiobiotin is a long-chain, amine-reactive labeling reagent. Its biotin-like group can be eluted from streptavidin, making it suitable for purification or labeling and purification of cell surface proteins.

EZ-Link Sulfo-NHS-LC-Desthiobiotin Features:

•Desthiobiotin—Biotin analogs can be easily synthesized from streptomycin Elutes in avidin, an ideal feature for affinity purification applications
•Protein Labeling—Label antibodies or proteins in purified or mixed samples for use with streptomycin affinity purification Achieving high recovery rates in sedimentation assays with peptide beads
•Cell surface markers—negatively charged reagents do not permeate cell membranes and therefore only modify surface proteins of whole cells
• Amine reactivity—Reacts with primary amines (-NH2) such as lysine side chains or amino termini of polypeptides
•Soluble—Reacts with common NHS esters The compound's charged sulfo-NHS group increases the water solubility of the reagent

Sulfo-NHS-LC-Desthiobiotin is a variant of biotin that is activated to have a long chain (LC ) sulfo-NHS ester of the spacer arm to covalently label proteins or other molecules with primary amines (-NH2) via the desthiobiotin group. Desthiobiotin tags bind streptavidin and other biotin-binding proteins with high specificity but tend to elute under mild conditions (i.e., in a manner that is competitively displaced by conventional free biotin). Therefore, this reagent is Sulfo-NHS-LC-Biotin (Product No. 21335), a technique where non-denaturing elution of tagged proteins is required. The negatively charged Sulfo-NHS group prevents the reagent from penetrating cell membranes; therefore, Sulfo-NHS-LC-desthiobiotin can be used to limit labeling of cell surface proteins in intact cells. No-Weigh format (5 x 1 mg resealable vials) eliminates the difficulty of weighing small amounts of reagents and protects unused reagents from hydrolysis as much as possible.

Comparison between desthiobiotin and biotin
Desthiobiotin is a monocyclic, sulfur-free biotin analog , the specificity of binding to strepavidin is almost the same as that of biotin, but the affinity is lower than biotin (1/Kd is 10¹¹ M and 10¹⁵ respectively M). Therefore, desthiobiotinylated bait proteins and their interacting partners can be easily and specifically washed from streptomycin affinity resins using mild conditions based on competitive displacement with free biotin. Come out. For sedimentation assays using biological samples, this soft release characteristic of desthiobiotin also helps to minimize co-purification of endogenous biotinylated molecules when elution of target protein complexes with free biotin Still bound to strepavidin. The modified avidin-biotin affinity system also eliminates the need for harsh elution conditions that may separate the complex and/or damage the target protein or cells. Desthiobiotin technology is ideal for working with native or recombinant proteins that do not express fusion tags or for isolating capture proteins under native conditions, such as when targeting intact cells or cell surface proteins.

Labeling using NHS ester reagent
Biotin'sN-Hydroxysulfosuccinate Imide (NHS) esteris the most commonly used biotinylation reagent. NHS-activated biotin reacts efficiently with primary amine groups (-NH2) in alkaline buffers to form stable amide bonds. Proteins typically have several primary amines that can be targeted for labeling, including the side chain of a lysine (K) residue and the N-terminus of each polypeptide. Most sulfo-NHS esters are directly water soluble but not membrane permeable. The unbleached plain NHS ester is less soluble in aqueous buffers but is membrane permeable.