An Electrochemical Biosensor Based On Aunp-Modified Gold Electrodes For Selective Determination Of Serum Levels Of Osteocalcin

Introduction

The concept of using immunological components as sensing agents was first described within an immunoassay in 1959 [1]. Since then, biomarker detection by antibody (Ab) conjugates has been a widely used disease diagnostic tool, with enzyme-linked immunosorbent assay (ELISA) being a gold standard even for comparison against all newly developed immunoassays and immunosensors [2]. The widespread use of Abs regardless of their high cost, variable affinity, and short shelf life arises from their exceptional specificity and sensitivity toward their binding partners, i.e.the antigens (Ag) [3].

While immunoassays have an extensive application in clinical and research areas, the immunosensor concept is yet to be implemented in routine diagnostics [4], [5]. There are certain issues in this regard, the most important of which is the coupling chemistry of proteins on surfaces, which is critical for optimal functioning. In other words, the optimal orientation of Ab molecules on the surface is of great importance for selective and sensitive detection ofAg in a reproducible way [6].

Immobilization methods vary largely based on the surface and protein properties. Ideally, as mentioned earlier, the protein conformation should remain intact during the immobilization process so that the protein retains its function. Moreover, the active sites of Ab should be accessible to reaction partners in order to have a high-performance, reproducible assay [7].

As for specialized detection methods such as electrochemical analysis, the protein is mainly immobilized on noble metal surfaces [8], [9]. One of the most commonly used surfaces in this regard is gold [10]. Gold nanoparticles are widely recognized as ideal supports for the fabrication of electrochemical sensors, mainly owing to their superior stability, high surface-to-volume ratio, its protein-friendly environment, capacity for surface modification and complete recovery in biochemical redox processes [11].

This article reports our experiments on immobilizing corresponding antibody on gold electrodes for electrochemical measurement of Osteocalcin (Oc), a bone turnover marker, levels in human serum. A step-wise protocol to develop the sensor is also included.