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Interaction with Streptavidin In order to calculate the number of binding sites of streptavidin (Sav), to a solution of Sav (5 µM, 200 µL), in phosphate buffer (10 mM, pH 7.4), a solution of HABA (2- (4-hydroxyphenylazo) benzoic acid, 2 mM) was added at regular intervals (7 min, 40 µL of total added volume), measuring the absorbance at 500 nm [24]