An antibody-based platform for melatonin quantification

Abstract

Melatonin, the 'chemical signal of darkness', is responsible to regulate biological rhythms and different physiological processes. It is mainly produced by the pineal gland as a hormone in a rhythmic daily basis, but it may also be synthesized by other tissues, such as immune cells, under inflammatory conditions. Its abnormal circulating levels have been related to several diseases such as type 2 diabetes, Alzheimer's disease and some types of cancer. Currently, melatonin is exclusively quantified by ELISA or radioimmunoassays, which although are very sensitive techniques and present low detection limits, usually require specialized personal and equipment, restricting the tests to a limited number of patients. To overcome such limitations, we developed a novel easy-to-use electrochemical immunosensor for rapid melatonin quantification. Anti-melatonin antibodies were immobilized into Indium tin oxide (ITO) platforms using (3-Aminopropyl)triethoxysilane (APTES), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS) crosslinkers. The platforms were assayed with synthetic and biologically-present melatonin containing samples. The developed device displayed a linear response in the concentration range from 0.75 to 7.5 μmol/L and a limit of detection of 0.175 μmol/L using Electrochemical Impedance Spectroscopy (EIS) (R² = 0.989) and 0.513 μmol/L using Cyclic Voltammetry (CV) (R² = 0.953) for synthetic melatonin. Furthermore, the sensors exhibited a good stability and reproducibility (3.45% and 2.87% for EIS and CV, respectively, n = 3), maintaining adequate response even after 30 days of assembly. On biologically-present melatonin-containing samples the device displayed a similar performance when compared to ELISA technique (deviation of 13.31%). We expect that the developed device contributes significantly to the medical area allowing precise and complete diagnosis of the diseases related to abnormal levels of melatonin.

Keywords: Antibody; Electrochemical biosensor; Electrochemical impedance spectroscopy; Hormone; Melatonin.