We have developed an effective competitive electrochemical immunosensor assay based on hapten-grafted programmed probe (HGPP) as a corecognition element for highly sensitive and selective detection of acetamiprid. Starting with the synthesis of hapten, HGPP was prepared using carboxyl group in the hapten and amino group in the 5' end of the programmed probe through covalent conjugation. Acetamiprid present in samples competes with HGPP to bind with capture antibody on the electrodes by specific recognition interaction. Methylene blue probe (MBP) was used as the electrochemical redox probe to capture the hybridized HGPP on the electrodes. The competitive reaction changes in accordance with the quantity of the target acetamiprid in the sample, as the amounts of the hybridized HGPP and the immobilized antibody are constant, i.e., the more acetamiprid samples are added, the less MBP is combined on the electrodes. In the optimal conditions, thus, biosensor output showed a linear relationship from 5 to 105 ng L-1 for the acetamiprid assay with a detecting limit of 3.2 ng L-1. The biosensor was successful in quantifying the amount of acetamiprid in spiked strawberry and cabbage extracts. This competitive immunosensor assay represents a rapid and sensitive technology for acetamiprid assay or other small molecule targets in food.
Keywords: acetamiprid; competitive immunoassay; electrochemical biosensor; hapten; pesticide residues.