The Research Of Electrochemical Biosensor For Antibiotics Determination Based On Body Conformation Chang Of Aptamer And Nuclease Amplification Technology

Nowadays,the abuse of antibiotics have caused the its residue in animal-derived foods such as eggs,milk,meet etc.Long-term intake of food containing the residue of antibiotics will resulted in the its accumulate and resistant in human body,being harm to human health.The residue of antibiotics has always been the one of the most crucial sources of pollution.Thus,it is urgent to establish a method to detect the antibiotics residue efficiently and rapidly.Aptamers have been an common tool in detection methods,due to its affinity and specificity of aptamer to target molecule.The nucleases have been widely applied in various an analysis methods with the advantages of stability,high efficiency.In this paper,we established two electrochemical biosensors based on the body conformation of aptamer combining with the using of nucleases to detect antibiotics,selecting kanamycin as the model analyte.With a wide linear range and a low detection limit,the biosenors showed excellent accuracy in real sample analysis.In the first approach,we established a electrochemical biosensor based on the body conformation of aptamer combining with the lambda exonuclease(? exo)and endonuclease(Nb.BbvCI)assisted recycle amplification strategy to detect kanamycin specifically and ultrasensitively.In the homogeneous solution,the skillfully designed arched probe contains kanamycin aptamer and a universal primer sequence,with the recognition of aptamer with target antibiotics,the primer was released and opened the hairpin probe 1(HAP1),forming the duplex structure.The primer opened the next HAP1,a recycle of primer was achieved,at the same time,plenty of triggers produced,with the digestion of ? exo.The triggers combined with the loop section of hairpin probe 2(HAP2)modified on the electrode which containing the restriction enzyme cutting site forming a duplex structure.With the recognition and cleavage of Nb.BbvCI,the trigger recycled,and the G-rich sequence in the HAP2 was completely exposed.In the presence of K+ and hemin,the G-quadruplex/hemin complex as horseradish peroxidase(HRP)-mimicking DNAzyme was formed.The quantitative determination of kanamycin was achieved by measuring the electrochemical signal transduction due to the reduction of H2O2.The sensitivity of sonsor was obviously improvedafter the nucleases assisted recycles.The results will detect kanamycin with the wide linear range from 1 pM to 10 nM,with a low detection limit as 0.5 pM.In the second approach,we established a signal-on electrochemical biosensors based on exonuclease III-assisted to detect kanamycin.The hybridization of Helper and 5'-methylene blue(MB)labeled HAP2,make the MB far from the surface of electrode,hindering the electron transferring.In the homogeneous solution.When the skillfully designed arched probe,which contained kanamycin aptamer and a universal primer sequence,was challenged with target,the recognition of aptamer with target antibiotics,the released primer opened the hairpin probe 1(HAP1).With the assistance of Exonuclease III(Exo ?),the primer was recycled and opened the next HAP1.At the same time,the produced trigger replaced the Helper from the duplex structure.HAP2 formed a “close” probe structure,confining MB close to the electrode surface for efficient electron transfer.The quantitative determination of kanamycin was achieved by measuring the increased electrochemical signal.The results will detect kanamycin with the wide linear range from 1 pM to 10 nM,with a low detection limit as 0.76 pM.