Development of a very sensitive electrochemical immunosensor for the determination of 17β-estradiol in bovine serum samples

Abstract

This study describes the development of a very sensitive electrochemical immunosensor (EI) for the determination of 17β-estradiol (17β-E). The novelty of this immunosensor is that the detection of 17β-E is carried out without sample pretreatment and unlabeled neither the antigen nor the antibody. Very good results in terms of sensitivity, kinetics, and working range are obtained. The immunosensor was constructed by immobilization of the anti-17β-E monoclonal antibody (mAbE) on a gold disk electrode modified with gold nanoparticles on a cysteamine self-assembled monolayer (AuNP-cys-Au disk). Bovine serum samples were spiked with known amounts of 17β-E and incubated on mAbE-AuNP-cys-Au disk electrodes. Then, the EI was transferred to pH 5.00 citrate buffer solutions, containing horseradish peroxidase (HRP), pyrocatechol (H2Q), and H2O2 at given concentrations. The 17β-E and H2Q, both enzyme co-substrates, react with HRP. The HRP, which did not react with 17β-E, in the presence of H2O2 catalyzes the oxidation of H2Q to o-benzoquinone (Q). The back electrochemical reduction of Q to H2Q was detected on the modified gold electrode surface by square wave voltammetry. The electrochemical signal was proportional to the amount of H2Q that reacts with the enzyme, and inversely proportional to the amount of 17β-E presents in the bovine serum samples. The EI showed a linear range from 0.54 to 1.36 × 104 pg mL-1. The limit of detection (LOD) was 0.84 pg mL-1. Recovery percentages were very good, with values of 99.7, 106, and 105% for 10, 50 and 100 pg mL-1, respectively. This EI is an attractive tool for the 17β-E determination in bovine serum samples.