Single-stranded DNA binding protein coupled aptasensor with carbon-gold nanoparticle amplification for marine toxins detection assisted by a miniaturized absorbance reader.

ABSTRACT

Okadaic acid (OA), one of the most widely distributed marine toxins worldwide poses a severe threat to human health. Previous sensing methods for OA detection are usually based on antigen-antibody binding mechanism. However, the drawbacks of antibodies especially the enzyme-labeled antibodies, such as the harsh storage condition and high cost, lead to significant challenges to OA detection in biological samples. To overcome these limitations, a single-stranded DNA binding protein (SSB) coupled aptasensor was developed for OA detection. SSB was incubated on the microplate as a substitute for conventional OA-protein conjugations. Carbon-gold nanoparticles were synthesized and labeled with horseradish peroxidase and thiol-modified aptamers to obtain a capture probe (CGNs@HRP-Apt) instead of the enzyme-labeled antibody for signal amplification. OA and SSB competed to bind with limited aptamers on CGNs@HRP-Apt probes followed by colorimetric assay to obtain the optical signals correlated to OA concentration. To achieve on-site detection, a miniaturized and multichannel absorbance reader (Smart-plate reader) was self-designed with full automation for OA detection. Utilizing the SSB coupled aptasensor and the Smart-plate reader, our approach enables cost-effective and on-site OA sensing with a detection range of 2.5-80 ppb and an ultra-low limit of detection of 0.68 ppb. Moreover, novel OA detection kits based on the SSB coupled aptasensor were prepared which can effectively reduce the cost by 15 times lower than that of commercial ELISA kits. Therefore, the developed platform provides a favorable and promising avenue for marine toxin detection in aquaculture and food safety.