Electrochemical Detection of Harmful Algae by Means of a Sandwich Hybridization Assay on an Electrode Surface

We report on the development of a sandwich hybridization assay on an electrode surface-based sensor device for the detection of toxic algae. This DNA (rRNA)-based sensor uses an electrochemical detection of a target sequence by means of its hybridization to a capture probe, with an attachment label at its 5′ end that binds it to the electrode surface, and a signal probe with a DIG detection label at the 3′ end. The thiol (biotin)-labelled capture probe is immobilized onto gold (carbon, graphite or carbon nanotubes) screen printed electrodes. Synthetic positive control DNA or real rRNA isolated from algal cells is applied to the sensor and allowed to hybridize to the capture probe. A signal probe with a digoxigenin label is then hybridized to this construct, followed by an anti-DIG-antibody coupled to horseradish peroxidase (HRP) and a substrate. The electrical signal obtained from the redox reaction is proportional to the amount of DNA (rRNA) applied to the biosensor, which is in turn proportional to the number of cells harvested when applied to real samples. The optimized hybridization conditions and steps of the fabrication process are presented here. The biosensor has been used to detect different algal species, such as Prymnesium parvum, Gymnodinium catenatum, Pseudo-nitzschia australis, Alexandrium ostenfeldii and Alexandrium minutum , by immobilizing their genetic material over the different transducer platforms.