Ratiometric absorbance and fluorescence dual model immunoassay for detecting ochratoxin a based on porphyrin metalation

In this work, a porphyrin metalation-based ratiometric absorbance and fluorescence dual model immunoassay was proposed to detect ochratoxin A (OTA). 5,10,15,20-tetrakis(1-methyl-4-pyridinio) porphyrin (TMPyP) was pink and had a strong fluorescence, upon coordination with Hg(II), its fluorescence was quenched and the color became green. The alkaline phosphatase can catalyze the dephosphorylation of ascorbic acid 2-phosphate to produce ascorbic acid, which can reduce the coordinated Hg(II) to Hg(0) and then dissociated from TMPyP, its fluorescence was recovered. Meanwhile, the color changed from green to light pink, which can be identified by naked eye for semi-quantitative detection. The linear ranges of ratiometric absorbance and fluorescence model were 0–6.0 ng/mL and 0.1–6.0 ng/mL, respectively. The absorbance and fluorescence signals produced by porphyrin metalation can mutually verify to improve the accuracy of detection results. Besides, the ultra-sensitivity and high selectivity demonstrated this method was a powerful tool for trace OTA detection.