Cascade-amplification-based electrochemical detection of Akashiwo sanguinea at pre-outbreak stage.

Red tide events caused by Akashiwo sanguinea (A. sanguinea) pose a significant threat to ecosystems. However, studies that offer promising approaches for portable and onsite detection with precise identification of A. sanguinea remain insufficient. In this study, we developed an electrochemical biosensor (E-biosensor) for detecting A. sanguinea combined with cascade amplification strategies, termed TDW-E-biosensor. A predictive relationship was also established to predict algal cell density based on electrochemical signals. The experiment results showed that the TDW-E-biosensor was successfully applied for detecting A. sanguinea at the pre-outbreak stage and demonstrated excellent analytical performance, showing a low limit of detection (LOD) of 0.0676 fM and quantitation (LOQ) of 0.102 fM for the three-electrode system, and a low LOD of 6.873 fg μL^-1 and LOQ of 20.460 fg μL^-1 for the portable system. The accuracy of the TDW-E-biosensor was validated through comparison with droplet digital PCR (ddPCR) and Bland-Altman analysis, demonstrating a high level of agreement (a mean difference of 0.132 and a standard deviation of 0.184). The reliability of the predictive relationship was evidenced by controlled laboratory experiments and Bland-Altman analysis. The developed TDW-E-biosensor provides an innovative and promising tool for early warning efforts regarding harmful algae.