The Study On Immunosensor Based On Ochratoxin A Phage Peptide Mimotope

Ochratoxin A(OTA)is a secondary metabolite produced by Penicillium spp and Aspergillus spp,and has a strong carcinogenicity to humans and animals.Therefore,countries have generally strengthened regulation of OTA.As an important means of analysis,immunosensor has a pivotal position in the OTA detection.In this paper,modify the gold nanoparticles(GNPs)and PEG polymers(Mal-PEG-NH2)on the gold electrode(GE)surface,obtained the functionalized PEG polymer brushes.The OTA immunosensor based on the principle of immunological competition was established by binded functionalized PEG polymer brushes covalently immobilized the anti-OTA-McAbof on it,and binded specifically OTA phage mimotopes.1 ELISA based on OTA phage peptides mimotopesTwo ELISAs were established by using of OTA phage peptide mimotopes and OTA artificial antigen respectively.The linear range of he conventional ELISA was0.20 ~ 13.52 ng/mL and the half inhibition concentration(IC50)was 1.66 ng/mL.Phage-ELISA linear was from 0.01 to 0.57 ng/m L and the IC50 was 0.09 ng/m L.2 Based on OTA phage peptides mimotopoes to establish immunosensor2.1 GNPs-immunosensorIn this work,3-mercaptopropionic acid(MPA)was immobilized on GNPs modified gold electrode surface by covalent bond and it can further used for fixing anti-OTA-McAb.Thus the GNPs-immunosensor was prepared,the working range of this sensor was 0.15~1.84 pg/mL,the IC50 was 0.53 pg/mL,and detection limit of20.48 fg/mL.2.2 GNPs-Mal-PEG-NH2-immunosensorGNPs was electrodeposited on gold electrode surface by cyclic voltammetry(CV),then the sensor was fabricated by layer-by-layer coating MPA ? Cys and Mal-PEG-NH2 on the working electrode,which can improve the surface area to capture a large number of anti-OTA-McAb.The IC50 of this sensor was 81.85 fg/m L,the linear range was 9.51~703.84 fg/mL with a detection limit of 2.05 fg/mL.Compared with GNPs-immunosensor,its sensitivity was nearly 10-fold higher than the GNPs-immunosensor.The prepared sensors were cross reacted with AFB1 ?DON?ZEN and FB1,and the results showed that the sensor had good specificity;Using beer and corn as a model sample,acceptable recovery of 99.33%~107.10%,89.61%~107.40%.3 The beer sample detection25 samples of beer were detected by using the prepared GNPs-Mal-PEG-NH2 immunosensor,commercially available ELISA kits and high performance liquid chromatography respectively,samples contaminated by OTA have not been found.