A novel electrochemical piezoelectric label free immunosensor for aflatoxin B1 detection in groundnut

• Highly ordered 4-ATP as nanowire for immune sensing.
• Developed EQCM immunosensor is more sensitive than QCM.
• The kinetic study proves it is a diffusion controlled process.
• The immunosensor can detect AFB1 in contaminated groundnuts samples.

An electrochemical quartz crystal microbalance (EQCM) based label-free immunosensor has been developed for the quantitative detection of aflatoxin B1 (AFB1) in groundnut. The gold (Au) coated quartz crystal (6 MHz) functionalized with self-assembled monolayer (SAM) of 4-aminothiophenol (4-ATP) has been utilized to immobilize anti-aflatoxin antibodies (aAFB1). The quartz crystal microbalance QCM frequency and EQCM cyclic voltammetry measurements have been used to optimize the deposition of SAM layer on Au electrode (4-ATP/Au), antibody immobilization (aAFB1/4-ATP/Au), bovin serum albumin (BSA) adsorption (BSA/aAFB1/4-ATP/Au) and immuno interaction (AFB1/BSA/aAFB1/4-ATP/Au). Interestingly, the EQCM studies conducted on this immunoelectrode provide wider linear range of 0.1–4.0 ng mL−1 and higher sensitivity than those obtained using under identical condition. The high association constant of 9.4 × 10−2 ng mL−1 of the EQCM immunosensor can be attributed to molecularly oriented self assembled monolayer of 4-ATP on QCM gold electrode that acts as a nanowire between aAFB1 and electrode through amide linkage formed by the NH2 group of 4-ATP and COOH group of Fc part of aAFB1. This linkage also helps to project paratope part of aAFB1 to interact with AFB1. The developed label free EQCM based immunosensor was tested for the AFB1 detection in the extract of contaminated groundnut and the results were compared with Liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. The comparative results demonstrate the suitability of this sensor for routine screening of groundnut and the percent error is found to be ±12%.