A novel colorimetric immunoassay for sensitive monitoring of ochratoxin A based on an enzyme-controlled citrate-iron() chelating system

Biotoxins present in foods represent a major source of contamination. Although different analytical methods have been developed, a new detection method for identification and quantification of these toxins is necessary for extending their application sphere. Therefore, in this study, an enzyme-controlled citrate-iron( iii ) chelating system was used to build a novel colorimetric immunoassay sensing platform for detection of ochratoxin A (OTA). The experimental signal was derived from the reaction of iron( iii ) and 3,3′,5,5′-tetramethylbenzidine (TMB). However, the addition of citrate could combine with iron( iii ) rapidly, thereby hindering the reaction of iron( iii ) and TMB for production of the signal ( i.e. , colorless to blue). Additionally, hydrogen peroxide (H 2 O 2 ), which could be produced from glucose in the presence of glucose oxidase (GOx), could affect the binding capacity of citrate and iron( iii ), thereby releasing iron( iii ) to react with TMB. That is to say, different amounts of GOx could regulate the intensity of the signal. Based on these findings, magnetic bead (MB)-based OTA competed with dissociative OTA (added) for glucose oxidase (GOx)-labelled anti-OTA. As the dissociative OTA concentration increased, the amount of GOx-labelled anti-OTA on MB conveniently reduced and the absorbance decreased. The method exhibited a good linear relationship as the concentration of the target (OTA) increased from 0.005 to 5 ng mL −1 ; the detection limit was 4.2 pg mL −1 , as estimated based on the 3S blank level under optimal conditions. The specificity and feasibility for a real sample of the colorimetric immunoassay was acceptable, respectively. The current results provided important insights into the development of citrate-iron( iii ) chelating system-based biological detection methods and colorimetric immunoassays. Schematic illustration of an enzyme-controlled citrate-iron( iii ) chelating system-based colorimetric immunoassay for sensitive determination of ochratoxin A.