Colorimetric immunoassay of furazolidone metabolites based on iron-copper bimetallic organic framework nanoenzyme.

Based on the peroxidase activity of Cu-hemin metal-organic framework (Cu-hemin MOF) nanozyme, a colorimetric enzyme-linked immunosensor was developed for the detection of furazolidone (FZD). Cu-hemin MOF is a bimetallic nanozyme that exhibited a stronger catalytic effect compared with single-metal organic framework nanoenzymes. Cu-hemin-MOF catalyzes hydrogen peroxide (H2O2) to produce hydroxyl radicals (•OH), which oxidizes the chromogenic substrate 3,3',5,5'-tetramethylbenzidine (TMB) to blue oxidized TMB (oxTMB). The absorbance change is at 650 nm. The content of AOZ in animal food can be quickly and accurately determined by changes in absorbance. The linear range of the colorimetric biosensor for detecting FZD was 0.01 ~ 62.52 ng/mL, and the limit of detection was as low as 0.01 ng/mL. The recovery of spikes samples was in the range 94.2-108.0 % and reproducibility was less than 4.8%. In addition, the cross-reaction rate was less than 0.1% when detecting other metabolites except AOZ, indicating that the sensor has good applicability and specificity. This study not only provides a better understanding of the relationship between the dispersion of nanoenzymes and enzyme-like activity but also offers a general method for detecting antibiotics using the nanoenzyme colorimetric method.