biostudies-literatureZhang ZNational Natural Science Foundation of ChinaNatural Science Foundation of Hainan ProvinceKey Research and Development Project of Hainan Province21760-21766https://www.ebi.ac.uk/biostudies/studies/S-EPMC9034093biostudies-literatureUnknown11(35)Ochratoxin A (OTA) is a common food contaminant with multiple toxicities and thus rapid and accurate detection of OTA is indispensable to minimize the threat of OTA to public health. Herein a novel enzyme cascade-amplified immunoassay (ECAIA) based on the mutated nanobody-alkaline phosphatase fusion (mNb-AP) and MnO₂ nanosheets was established for detecting OTA in coffee. The detection principle is that the dual functional mNb-AP could specifically recognize OTA and dephosphorylate the ascorbic acid-2-phosphate (AAP) into ascorbic acid (AA), and the MnO₂ nanosheets mimicking the oxidase could be reduced by AA into Mn²⁺ and catalyze the 3,3',5,5'-tetramethyl benzidine into blue oxidized product for quantification. Using the optimal conditions, the ECAIA coulRSC advancesEnzyme cascade-amplified immunoassay based on the nanobody-alkaline phosphatase fusion and MnO₂ nanosheets for the detection of ochratoxin A in coffee.PMC903409331901800320RC509317604932019RC119ZDYF2020157Liu XChen QSu BCao HZhang ZHe ZXu HZhou YSun ZfalseEnzyme cascade-amplified immunoassay based on the nanobody-alkaline phosphatase fusion and MnO₂ nanosheets for the detection of ochratoxin A in coffee.Ochratoxin A (OTA) is a common food contaminant with multiple toxicities and thus rapid and accurate detection of OTA is indispensable to minimize the threat of OTA to public health. Herein a novel enzyme cascade-amplified immunoassay (ECAIA) based on the mutated nanobody-alkaline phosphatase fusion (mNb-AP) and MnO₂ nanosheets was established for detecting OTA in coffee. The detection principle is that the dual functional mNb-AP could specifically recognize OTA and dephosphorylate the ascorbic acid-2-phosphate (AAP) into ascorbic acid (AA), and the MnO₂ nanosheets mimicking the oxidase could be reduced by AA into Mn²⁺ and catalyze the 3,3',5,5'-tetramethyl benzidine into blue oxidized product for quantification. Using the optimal conditions, the ECAIA coul2021-01-01T00:00:00Z2021 Jun2025-04-22T01:39:56.193Z2025-04-05T20:06:19.856ZS-EPMC90340933547880910.1039/d1ra03615g