HOOK™ BiotinQuant Kit, 20 Assays

Protocol

• A kit for estimation of biotin conjugation
• Biotin labeling is measured using HABA [4'-hydroxyazobenzene-2-carboxylic acid] dye.
• HABA binds with avidin at the biotin binding site.
• The HABAavidin complex produces a characteristic color that absorbs at 500nm. (ε = 34,000 M-1 cm-1 expressed as per mole of HABA bound).
• In the presence of biotin or biotinylated agents, these agents compete with the HABA for the binding sites.
• Since biotins have a greater affinity for the binding sites it results in the displacement of HABA from the avidin binding sites and proportionate reduction of absorbance.
• Hence, the amount of biotin present in the solution can be determined by measuring the reduction in absorbance at 500nm.