Bifunctional M13 Phage as Enzyme Container for the Reinforced Colorimetric-Photothermal Dual-Modal Sensing of Ochratoxin A

Point of care (POC) methods without expensive instruments and special technicians are greatly needed for high-throughput analysis of mycotoxins. In comparison, the most widely used screening method of the conventional enzyme-linked immunosorbent assay (ELISA) confronts low sensitivity and harmful competing antigens. Herein, we develop a plasmonic-photothermal ELISA that allows precise readout by color-temperature dual-modal signals based on enzymatic reaction-induced AuNP aggregation for highly sensitive detection of ochratoxin A (OTA). The bifunctional M13 phage carrying OTA that mimics the mimotope on the end of p3 proteins and abundant biotin molecules on the major p8 proteins is adopted as an eco-friendly competing antigen and enzyme container for amplifying the signal intensity. Under optimal conditions, both colorimetric and photothermal signals enable good dynamic linearity for quantitative OTA detection with the limits of detection at 12.1 and 8.6 pg mL(-1), respectively. Additionally, the proposed ELISA was adapted to visual determination with a cutoff limit of 78 pg mL(-1) according to a vivid color change from deep blue to red. The recoveries of OTA-spiked corn samples indicate the high accuracy and robustness of the proposed method. In conclusion, our proposed strategy provides a promising method for eco-friendly and sensitive POC screening of OTA. Moreover, it can be easily applied to other analytes by changing the involved specific mimotope sequence.

Automated summary

In order to achieve high-throughput analysis of mycotoxins, there is a great need for point-of-care (POC) methods that do not require expensive instruments or specialized technicians. The conventional enzyme-linked immunosorbent assay (ELISA), which is widely used for screening, has low sensitivity and is affected by competing antigens. In this study, we developed a plasmonic-photothermal ELISA that uses color-temperature dual-modal signals for highly sensitive detection of ochratoxin A (OTA), with the signal amplified by a bifunctional M13 phage. Under optimal conditions, the colorimetric and photothermal signals showed good dynamic linearity for quantitative OTA detection, with detection limits of 12.1 and 8.6 pg mL(-1), respectively. Additionally, the proposed ELISA could be visually determined with a cutoff limit of 78 pg mL(-1) based on a distinct color change from deep blue to red. The method also showed high accuracy and robustness when applied to OTA-spiked corn samples. Overall, this strategy provides a promising method for eco-friendly and sensitive POC screening of OTA, and can be easily adapted for other analytes by changing the specific mimotope sequence.