Biocytin hydrazide

The reactivity and use of biocytin-hydrazide is similar to that described for biotin-hydrazide in Section 3, this chapter. The following protocol for labeling glycoproteins at oxidized carbohydrate (galactose) sites is from Bayer and Wilchek (1992). 

Dissolve the glycoprotein to be labeled in 0.1 M sodium phosphate, 0.15 M NaCl, pH 7.4, containing 1 mM CaCl2 and 1 mM MgCl2 (labeling buffer), at a concentration of 1 mg/ml. 

To each ml of glycoprotein solution, add 30 pi of neuraminidase (1 unit/ml as supplied by Behringwerke AF), then 30 pi of galactose oxidase (previously dissolved at 100 units/ml in the labeling buffer of step 1), and finally 100 pi of the biocytin hydrazide solution. 

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Figure 11.12 Biocytin-hydrazide reacts with aldehyde-containing molecules to form hydrazone bonds.

Bayer, E.A., Ben-Hur, H., and Wilchek, M. (1988) Biocytin hydrazide—a selective label for sialic acids, galactose, and other sugars in glycoconjugates using avidin-biotin technology. Anal. Biochem. 170, 271-281. 

Prepare the labeling reagent stock solution. Biotin, biocytin, and desthiobiotin hydrazide derivatives are typically used at a labeling reagent antibody MR of 20-25. Biotin and desthiobiotin hydrazide stock solutions should be prepared at 10-20 mg/mL in DMSO or DMF while biocytin hydrazide can be dissolved in the reaction... 

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