ELECTROCHEMICAL EVALUATION OF AVIDIN-BIOTIN INTERACTION USING N-IODOACETYL-N-BIOTINYLHEXYLENEDIAMINE

The avidin-biotin assay was investigated by an electrochemical procedu re based on the chemical reaction between cysteine containing a thiol group and N-iodoacetyl-N-biotinylhexylenediamine (IB), In the presence of avidin and biotin this reaction, whereby the thiol group combines with IB, is controlled because IB has a biotin part; that is, avidin a nd biotin are detected indirectly by measurement of iodide ion or cyst eine. To achieve a high sensitivity of detection, Hg(II) was introduce d as a marker that interacts with cysteine and the oxidation peak from Hg(0) to HE(II) was measured. The sensitivity of detection of avidin was at the level of 10(-9) M. The relative standard deviation at 1 x 1 0(-8) M avidin was 4.8% (n = 5). On the other hand, a response curve t o detect biotin was obtained by the competitive reaction between IB an d biotin for the limited binding sites of avidin. The change in peak c urrent enables the detection of biotin at the level of 10(-9) M. This method has the advantage that it is not necessary to separate free IB from bound IB.