Rapid Determination Of Triazophos By Biomimetic Immunoassay Based On Molecularly Imprinted Recognition Material

Triazophos is a moderately toxic,broad-spectrum organophosphate insecticide that has been widely used in cereals,fruits and vegetables.Because of its good chemical stability and long half-life,triazophos is easy to remain in the environment,causing potential hazards to the environment and human health.Since the end of 2016,triazophos has been banned for use on vegetables by the Chinese Ministry of Agriculture.At present,the main detection techniques of triazophos include confirmation technologies and immunoassays,but these methods usually have many shortcomings,such as expensive instruments,long analysis time,difficult preparation of antibodies and so on.Therefore,it is of great practical significance to design and synthesize bionic recognition materials with strong specificity,good stability and low preparation cost,and to establish sensitive,simple,fast,stable and cheap detection methods.Based on molecular imprintied specific recognition materials,the following aspects have been carried out in this paper:molecular imprinting biomimetic enzyme-linked immunosorbent assay,biomimetic fluorescence immunosorbent assay and electrospinning molecular imprinting membrane chromatography.The details are as follows:?1?96-well molecular imprinted array membranes with good specificity were obtained with dummy template as design concept by bulk polymerization and surface imprinting.We synthesized the enzyme-labeled probe of triazophos and established a direct competitive biomimetic enzyme-linked immunosorbent assay?BELISA?.Under optimal conditions,the linear range of the method was0.001-10000?g/L with a good regression coefficient of 0.9770.The sensitivity(IC₅₀)and the limit of detection(LOD,IC₁₅)of BELISA were 428?g/L and 0.006?g/L,respectively.This method was performed to detect triazophos in cabbage and apple samples,and showed excellent recovery and relative standard deviations?RSDs?ranging from 70.5 to 119.8%and from 5.2 to 19.7%,respectively.The results correlated well with those obtained using high performance liquid chromatography-tandem mass spectrometry?R²=0.9927?.?2?Triazophos hapten CdSe/ZnS quantum dots labeled probes with ester bond coupling technique were synthesized and self-synthesized molecular imprinted 96-well array membranes as recognition elements were used in this experiment.Under the optimal conditions,a biomimetic fluorescence immunoassay based on molecular imprinting recognition technology was established.The assay showed an excellent linear response over the concentration ranges of 0.1-10000?g/L with a good coefficient of determination?R²=0.9817?.The sensitivity(IC₅₀)and limit of detection(LOD,expressed as IC₁₅)of the developed assay were 3.63 mg/L and 0.31?g/L,respectively.The applicability of the developed approach was tested for detecting triazophos in cabbage and apple samples.The method showed excellent recoveries?109.6-118.9%?and relative standard deviations?RSDs?between 9.9-19.5%.The obtained results correlated well with those obtained by LC-MS/MS?R²=0.9995?.The competitive assay using CdSe/ZnS QDs as fluorescence-labeled probe showed a good sensitivity,steady,fast response,and excellent anti-interference ability compared to conventional fluorescence-quenching methods.?3?In this experiment,nanocomposite films of nanofibers hybrided with molecularly imprinted microspheres were synthesized by electrospinning technology.Then,the THBu-IgG-FITC fluorescent probe was prepared,and nano-film chromatography strip with specific recognition of triazophos was preliminarily developed.Based on these,a new method for rapid detection of triazophos was established.We optimized the technical parameters and preparation conditions,such as spinning solution?type,solvent,concentration?,electrospinning conditions,surfactant,MIPs concentration,fluorescent probe concentration,competition mode,etc.The method showed an excellent linear response over the concentration ranges of 20-500?g/L with a good coefficient of determination?R²=0.9543?,and the limit of detection was 20?g/L.This method solves the technical bottleneck that molecularly imprinted microspheres can not be persistently fixed as a recognition element of bionic test strip,broadening the application scope of immunochromatographic test strip,and providing a new idea for subsequent researches.