HABA (4'-hydroxyazobenzene-2-carboxylic acid) is a dye that can be used for quantitation of biotin in a sample. When bound to avidin, the HABA-avidin complex has an absorbance at 500 nm (Ec = 34,000). Due to its higher affinity for avidin, biotin when introduced to a HABA-avidin solution will displace HABA and the absorption at 500 nm will decrease proportionately. Thus, we can utilize this method to estimate the moles of biotin in solution.

This calculator will use the change in max absorbance at 500 nm to estimate the concentration of biotin in solution with Beer's Law. Then it will return a mole-to-mole ratio of biotin to its linked protein (protein concentration must be known).

However, HABA-avidin based assays suffer from some inaccuracies that can be caused by a variety of factors like steric hindrance. For an alternative, see our product: Readiview Biotin Succinimidyl Ester