Thermo Scientific EZ-Link Hydrazide-PEG4-Desthiobiotin is a pegylated carbonyl reactive labeling reagent. Its biotin-like group can be eluted from streptavidin, making it useful for cell surface glycoprotein labeling and purification.

EZ-Link Hydrazide-PEG4-Desthiobiotin Features:

•Desthiobiotin—A biotin analog can be easily synthesized from streptomycin Elutes in avidin, an ideal feature for affinity purification applications
•Glycoprotein Labeling—labels glycosylated proteins at sialic acid residues for affinity purification in streptomycin Achieve high recoveries in sedimentation assays with cell surface microbeads
•Cell surface markers—label and isolate cell surface glycoproteins; reagent does not penetrate whole cell membranes
•Aldehyde reactivity —Reacts with aldehydes formed by glycosyl periodate oxidation
•Hydrazide Activation—In buffers (such as sodium acetate) at pH 4 to 6 The reaction is carried out in
•PEGylation—the spacer arm contains a hydrophilic 4-unit polyethylene glycol (PEG) group
•Enhanced solubility —PEGylation makes the biotinylated molecule water-soluble, helping to prevent aggregation of biotinylated antibodies stored in solution

This desthiobiotin variant of biotin is activated and can label large Molecular periodate oxidizes carbohydrates. Hydrazide groups are coupled to carbonyl groups (aldehydes and ketones), such as those produced by sialic acid and other sugar components of glycoproteoglycans after treatment with 1 to 10 mM sodium periodate (NaIO4). 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 used in avidin-biotin technology hydrazide-PEG4-biotin (Product No. 21360) by which nondenaturing elution of tagged proteins is expected. Hydrophilicity 4 unitsPolyethylene glycol (PEG) spacers enhance solubility and reduce aggregation of tagged proteins stored in solution. The PEG fragment also increases the length and flexibility of the spacer arm, thereby minimizing steric hindrance when binding to the avidin molecule. No-Weigh format (5 x 1 mg resealable vials) eliminates the difficulty of weighing small amounts of reagents and protects unused reagents from degradation as much as possible.

Desthiobiotin and biotin
Desthiobiotin is a monocyclic, sulfur-free biotin analogue. The binding specificity of streptavidin is almost the same as that of biotin, but the affinity is lower than that of biotin (1/Kd = 10^11 vs. 10^15M, respectively) [1-2]. 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.

Use hydrazide reagent for labeling
Hydrazides and alkoxyamines are two typesCarbonyl reactive group types . Hydrazide (–NH-NH2) reacts specifically with the aldehyde group under slightly acidic conditions to form a hydrazone bond; use Sodium cyanoborohydride (Product No. 44892)can further reduce it to a stable secondary amine bond. The reaction is more efficient in the presence of aniline (Product No. 88944). Alternatively, you can use EDC carbodiimide chemistryCouples hydrazides with carboxylic acids.

Sodium periodate (Product No. 20504) can be used to pass the sugar glycol group Partial oxidation generates reactive aldehyde groups in glycoproteins and other polysaccharide compounds. Sialic acid residues are common components of protein glycosylation and are easily converted to aldehydes with 1 mM NaIO4.