| Excess dose of pesticide was a great threat to food safety and human health. The onsite and rapid detection of pesticide residues was an effective way to monitor the rational using of pesticides and improve the safety of agricultural products. Processing of sample of the traditional instrument analysis method was complex and the instrument needs professional personnel to operate. So the traditional method was not suitable for the rapid onsite detection. The biosensor for pesticide residue detection based on microelectrodes had the advantages of good selectivity, short detection time, high sensitivity, easy miniaturization and other advantages and it was the hot spots about rapid detection of pesticide residue on the home and abroad and it also can realize the onsite online detection of agricultural products.In this paper, combining microelectrodes with biosensor technology, we fabricated a biosensor based on the microelectrode. Parameters that affecting the aptasensor performance of different modification materials and methods were studied. Taking electrochemical impedance value as response signal and the pesticide chlorpyrifos as template, we constructed the biosensor based on microelectrodes and used it for the rapid detection of pesticide residues in vegetable samples. The specific research contents and results were as follows:1. The feasibility of using the interdigitated array microelectrodes to construct the immunosensor for the detection of pesticide residues was explored. The methods of fixing antibody on the microarray electrodes(physical adsorption, protein A immobilization and methanethiol(MTB) / gold nanoparticles(Au NPs) collaborative assembly and protein A immobilization method) were compared. The results showed that the MTB/Au NPs self assembly and protein A directional fixation method for antibody immobilization can effectively maintained the activity of the antibody and increased the antibody immobilization amounts and the immune response. Therefore, we chose this method constructed the immunosensor based on microarray electrodes. The results showed that there is a good linear correlation between the impedance difference and pesticide concentrations in the range from 0.05 ng/ml to 500 ng/ml. The regression equation was ΔR= 382.6 + 84.545 lg C(ng/m L), the correlation coefficient was 0.97131 and the minimum detection limit was 0.017 ng/ml. The immunosensor was applied to the detection of lettuce,Chinese cabbage and cucumber, respectively. The recoveries of the three levels of the threesamples were studied respectively, and the recovery rate was 81.3- 99.7%.2. An immunosensor based on self assembly technique and protein A directed immobilization method and combined the microfluidic chip technology was applied to the detection of pesticide residues and realized the automatic feeding of samples. Using poly dimethyl diallyl ammonium chloride(PDDA) and nano gold nanoparticles(Au NPs)self-assembly to immobilize antibodies into microfluidic chip microchannel, solving the problem of blocking the micro fluidic channel. Under the optimum conditions, there is a good linear correlation between the impedance difference and pesticide concentrations in the range from 0.5 ng/ml to 500 ng/ml. The regression equation was ΔR = 163.0 + 70.000 lg C(ng/m L), and the correlation coefficient was 0.97454. The immunosensor was applied to the detection of cucumber, lettuce and cabbage, respectively. The three levels of the three samples were detected and the recovery rate was 75.2- 96.5%.3. The method of sensor based on the interdigitated array microelectrodes and aptamers was explored and it was used for the rapid detection of pesticide residues in vegetable samples. The sensor adopted aptamers instead of antibody based on the interdigitated array microelectrodes reduced the detection cost. Under the optimum conditions, there is a good linear correlation between the impedance difference and pesticide concentrations in the range from 0.05 ng/ml to 500 ng/ml. The regression equation was ΔR = 84.41+15.219 lg C(ng/m L), the correlation coefficient was 0.95995 and the minimum detection limit was 0.01 ng/m L. The aptasensor was used for the detection of pakchoi, cabbage and leeks respectively. The three levels of the three samples were detected and the recovery rate was 77.9- 100.2%. |
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