Dual-Readout Immunochromatographic Assay by Utilizing MnO2 Nanoflowers as the Unique Colorimetric/Chemiluminescent Probe

Manganese dioxide nanoflowers (MnO₂ NFs) were synthesized and used as a dual readout probe to develop a novel immunochromatographic test strip (ITS) for detecting pesticide residues using chlorpyrifos as the model analyte. MnO₂ NFs-labeled antibody for chlorpyrifos was employed as the signal tracer for conducting the ITS. After 10 min competitive immunoreaction, the tracer antibody was captured by the immobilized immunogen in the test strip, resulting in the captured MnO₂ NFs on test line. The captured MnO₂ NFs led to the appearance of brown color on the test line, which could be easily observed by the naked eye as a qualitative readout. Due to the very slight colorimetric difference of chlorpyrifos at trace concentrations, the semiquantitative readout by naked eyes could not meet the demand of quantitative analysis. MnO₂ NFs showed a significant effect on the luminol-H₂O₂ chemiluminescent (CL) system, and the CL signal driven by MnO₂ NFs were used to detect the trace concentration of chlorpyrifos quantitatively. 1,3-Diphenylisobenzofuran quenching studies and TMB-H₂O₂ coloration assays were conducted for studying the enhancing mechanism of MnO₂ NFs, which was based on the oxidant activity to decompose H₂O₂ for forming reactive oxygen species. Under optimal conditions, the linear range of chlorpyrifos was 0.1-50 ng/mL with a low detection limit of 0.033 ng/mL (S/N = 3). The reliability of the dual-readout ITS was successfully demonstrated by the application on traditional Chinese medicine and environmental water samples. Due to the simultaneous rapid-qualitative and sensitive-quantitative detection, the dual-readout protocol provides a promising strategy for rapid screening and field assay on various areas such as environmental monitoring and food safety.