| Pesticides are widely used in agriculture industry. They have contributed significant economic benefits to society. At the same time, widespread use of pesticides has created serious concerns regarding their effect on the environment and human health. The standard methods for identification and quantification of pesticides are generally based on chromatographic methods, such as gas chromatography (GC) or high-performance liquid chromatography (HPLC) coupled with mass spectroscopy (MS). These analytical techniques are very sensitive and reliable, but they are time-consuming and expensive. Moreover, they can only be performed by highly trained technicians and not convenient for on-site or in-field detection. Due to the great amount of pesticides currently being used, there is an increasing interest for developing rapid screening systems for their detection.Biosensors are analytical devices which tightly combine biorecognition elements with physical transducers for detection of the target compounds. Immunosensors are one kind of the biosensors, which have great potentials in pesticide detection. Due to the fact that organophosphorous pesticides (Ops) were most widely and commonly used, this research is focused on Ops pesticide residual detection by using electrochemical, piezoelectric and optical immunosensors.The main contents and results are summarized as follows.(1) The feasibility of electrochemical impedance spectra (EIS) used in the detection of pesticide residues was evaluated. It was shown that EIS in the process of electrode modification, antibody immobilization, and interaction of antibody-antigen were exactly consistent with the models of equivalent circuit. According to the Nyquist spectra, the change values of Ret, Rs, CPE, and Cdl were measured. It could be found that there were no significant changes in the elements of Rs and CPE, whereas the percentage changes of Ret and Cdl were notable. When antibody modified electrode was interacted with various concentrations of pesticide, a good relationship between Ret and concentrations of Ops pesticide was found ranging from 0.01μg/mL to 100μg/mL. The regression equation was y=250.54Ln(x)+1925.2, with the correlation coefficient of 0.9705. The low limit of detection was 2.3×10-2μg/mL. This electrochemical impedance immunosensor was evaluated in real water samples. The mean recovery rates with three levels of spiked samples were 85.3%,116.7% and 101.4%, respectively. The variation coefficients were 10.6%, 13.7%, and 14.5%, respectively. In addition, anti-pesticide antibodies were immobilized onto the magnetic beads, which were used to capture pesticide. After magnetic beads with captured pesticides were injected into microfluidics chips, different concentrations of pesticides can be easily differentiated in the Bode diagram.(2) A piezoelectric immunosensor coupled with a flow injection system was developed for detection of pesticide parathion. It was shown that in the range of 0.1-50μg/mL, there was a good corelative relationship between the concentration of pesticide parathion and the frequency shift, and the regression equation was y=10.827x+22.133 with coefficient of determination of 0.9862. The detection limit was 3.38×10-2μg/mL. Five methods for immobilization of the anti-parathion pesticides monoclonal antibody onto the surface of a gold electrode on quartz crystal were compared for effective capture of objective pesticides. According to sensitivity and repeatability, the effects of immobilization of protein A and silane methods were better than others. AFM images were also used in the evaluation of modification of electrodes.(3) Two competitive strategies, named antibody-immobilized method and antigen-immobilized method, were carried out to detect pesticide residues by using the quatz crystal microbalance. In the competitive antibody-immobilized method, the concentrations of immobilized antibodies were optimized. A correlative relationship between the concentration of objective pesticides and the frequency shift were found in the rang of 0.5 ng/mL～5μg/mL, when antibodies with dilution 1:30 and 80μg/mL artificial antigen were adopted. The regression equation was y= 11.555Ln(x)-158.1, with coefficient of determination of 0.9664. The low limit of detection was 6.19×10-2 ng/mL. In the competitive antigen-immobilized method, different concentrations of immobilized artificial antigens were optimized and performance of detection was evaluated. It was showed that effect of immobilization was optimal when the concentration of immobilized antigen was 200μg/mL and pH was 7.4. A good correlative relationship between the concentration and the frequency shift was obtained and the regression equation was y=15.711 Ln(x)-203.85 with coefficient of determination of 0.9862. The low limit of detection was 6.19×10-2 ng/mL. The immunosensor could be used repeatedly with good selectivity. In addition, cyclic voltammetry (CV) was employed to investigate the electrochemical character of immunosensor(4) A capillary based optical immunosensor was developed to detect pesticide triazophos, in which antigen-antibody reaction was coupled with biotin-avidin systems. Indirect immuno-competitive assays were used in this optical immunosensor. TMB substrate was catalyzed by HRP enzyme labeled complex, and the absorbance of product was measured at 655 nm. The concentration of objective pesticide was inverse proportion to values of absorbance. Calibration curves were constructed by plotting the percentage of A/Ao vs the logarithm of the concentration of pesticide triazophos. The regression equation was y=25.96x + 102.09 with coefficient of determination of 0.9874. Detection limit of 1.8 ng/mL was obtained for parathion by using Ao -2SD method, which was lower than stated MRL value. It accorded with requirement of residues analysis. The immnosensor was used in the detection of pool water, teas, and pears. The mean values of concentration of triazophos in three samples were 9.7 ng/mL, 6.4 ng/mL, and 3.5 ng/mL, respectively. Two levels of recovery test in three kinds of spiked samples were studied. Recovery rates of 103.7% and 104.8% were obtained for pool water. 78.7% and 85.6% were for tea. 103.5% and 105.6% were for pears. |
PDF Full Text Request