Development Of Immunoassays Based On Gold Nanomaterials For Imidacloprid

Neonicotinoid pesticides are a new class of pesticides with broad spectrum and low bioaccumulation.Imidacloprid is one of the most widely used neonicotinoid insecticides.As a low-dose systemic insecticide with significant insecticidal effects,imidacloprid has been used as a new generation of insecticides nationwide and is a good alternative to traditional insecticides such as organophosphorus,carbamate and pyrethroids.However,the widespread use of imidacloprid in agricultural production is bound to cause pesticide residues.Long-term consumption of grains,vegetables,fruits with pesticide residues,a certain amount of residual pesticides will accumulate in human body.Studies have shown that imidacloprid can affect the development of mammalian brain.It has neurotoxicity,reproductive toxicity and genotoxicity,large doses of imidacloprid can cause degenerative changes in the thymus,pancreas and bone marrow of animals.Long-term consumption of agricultural products containing imidacloprid residues can cause certain damage to human gastric mucosa.Therefore,it is of great significance to develop an efficient and practical method for detecting pesticide residues.On the basis of obtaining imidacloprid monoclonal antibody and imidacloprid coated antigen,three immunological detection methods were established based on antigen-antibody reaction,and the specific detection of imidacloprid in agricultural products was realized.The specific work of this paper is as follows:1、An ic-ELISA has been established and optimized.The linear regression eauation of the ic-ELISA was y=42.793x-65.32,with the correlation coefficient（R²）of 0.9967.The sensitivity(IC₁₀)and the linear range（15–85%inhibition）of the ic-ELISA were 0.06μg/L and 0.08 to 3.26μg/L respectively.the assay had almost no CR with other analogues（<1.00%）except for thiacloprid（8.48%）,acetamiprid（4.02%）and clothianidin（1.18%）,The recoveries of imidacloprid from spiked samples were ranged from 83.61 to 112.65%,these results showed a good correlation between the developed ELISA and commercial kit（r=0.9763）.2、Firstly,AuNPs-HRP was formed by AuNPs loaded with a large amount of HRP.Then LBL technology was used to encapsulate a large number of AuNPs-HRP into a capsule shape.Finally,nanocapsule probes were formed by coupling secondary antibodies on the surface of the capsules.Based on this probe,an detection method based on nanocapsule probe was established and optimized.The linear regression eauation of the assay was y=39.719x-31.238,with the correlation coefficient（R²）of0.9487.The sensitivity(IC₁₀)and the linear range（15–85%inhibition）were 0.01μg/L and 0.02 to 0.84μg/L respectively.The recoveries of imidacloprid from spiked samples were ranged from 80.34 to 120.84%,these results showed a good correlation between the developed assay and commercial kit（r=0.9869）.3、AuBPs was prepared by seed growth method.ALP catalyzes the 4-APP to produce 4-AP and phosphate ions.4-AP has reducibility,can reduce Ag⁺to Ag and deposit on the surface of AuBPs to cause changes in its LSPR properties（color change and shift of maximum absorbption peak wavelength）.This reaction was introduced into conventional ic-ELISA to establish p-ELISA based on silver deposition AuBPs.The linear regression eauation of the assay was y=-57.691x+262.22,with the correlation coefficient（R²）of 0.992.The linear range were0.781 to 25μg/L.The recoveries of imidacloprid from spiked samples were ranged from 76.82 to 124.17%,these results showed a good correlation between the developed assay and commercial kit（r=0.9740）.Our developed three detection immunoassays can meet maximun residue limits（MRLs）of imidacloprid in agricultural products in China.The recoveries of imidacloprid were in accordance with Association of Official Analytical Chemists（AOAC）standards.The developed three immunoassays are feasible for the detection of cereals,fruits and vegetables.