| The residues of kanamycin in animal-originated agricultural products,soil,and water bodies will have a serious impact on human health and the ecological environment.Therefore,strict monitoring of kanamycin residues in agricultural products and environmental samples should be conducted.However,traditional detection methods are time-consuming,complex,and require specific equipment or personnel,thus limits their application.The optical biosensor has the advantages of simple,convenient,fast response,low detection limit,good accuracy,high sensitivity and low cost.It has developed rapidly in the analysis field.Gold nanomaterials have unique optical properties,making them widely used in optical sensing.In the detection,no enzyme-assisted hybridization chain reaction(HCR)can achieve signal amplification at room temperature,so as to achieve the purpose of trace detection of the target.In this paper,a colorimetric method using gold nanomaterials as the signal conversion element and HCR technology as the signal amplification method was established,and the determination of kanamycin was achieved.The research content and results of this paper are as follows:(1)An optical biosensing method based on gold nanoparticles(AuNPs)and HCR-assisted amplification was established to detect kanamycin in milk.Firstly,HCR hairpin probe containing aptamer of kanamycin was designed and characterized by gel electrophoresis.Subsequently,the feasibility of the detection method was analyzed,and its key parameters were optimized:the optimal concentration of AuNPs was 6.5 nM,the optimal conjuncted time of the AuNPs and the HCR product was 30 min and the best measuring time was 40 min after salt addition.The optical biosensing method established in this study has good linear to kanamycin in the concentration of 2.14 to 40 ?M with a detection limit of 0.68 ?M and exhibits high selectivity.Kanamycin can be quantitatively or qualitatively detected by measuring absorbance changes and color changes.In addition,it was successful used in detection of kanamycin in milk samples.this work demonstrates that by changing the corresponding aptamer sequences and hairpin DNA probes,the established optical biosensing method can be extended to detect the residue of other small molecule substances in agricultural products,and it has practical potential for the safety of agricultural products and human health.(2)Based on the work of AuNPs,we proposed to introduced AuNRs with tunable optical properties to establish corresponding optical biosensing method for kanamycin detection.A preliminary study of the gold nanorod colorimetric method based on HCR amplification strategy was done for randomly selected sequence DNAR and the corresponding optical biosensing method was estabised for the first time.Firstly,the AuNRs with a ratio of 2.77 ± 0.30 were synthesized by the seed growth method,then the designed DNA sequences was characterized by gel electrophoresis.Meanwhile,the feasibility of the detection method was analyzed and the key parameters were optimized:the optimal concentration of HCR hairpin probe was 250 nM,and the best mearsuring time was 4 min after salt addition.The optical biosensing method established has good linear to DNAR in the concentration of 4.62 to 60 nM with a detection limit of 1.47 nM,and it exhibits extremely high selectivity which can distinguish single-base mismatched DNA.This proves that AuNRs can be used to establish optical biosensing methods based on HCR amplification strategy.By introducing the aptamer hairpin probe,this study can provide an effective method basis for highly sensitive detection of antibiotics(including Kana),mycotoxins and other substances.Moreover,It provides a promising application for the established AuNRs-based and HCR assisted optical biosensing method in sensing field and expands the potential scope of AuNRs in building biosensing method. |
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