Study On Preparation And Application Of Polyclonal Antibody To Quizalofop-p-ethyl

Immunoassay is an important tool for analysis and detection of pesticide residue and its metablital derivatives and other smell molecules. In this work, we have developed immunlolgical and instrumental assays for quizalofop-p-ethyl, a hapten was designed for quizalofop-p-ethyl, and then it was conjugated with different carrier proteins to synthesize the immunogens and coating antigen in different ways, CI-ELISA were established with corresponding polyclonal antibodies.What's more, we determined quizafolop-p-ethyl pesticide residues in water and soil.1. To produce a novel type of immunogen susceptible to raise antibodies, on the alkaline condition, quizalofop-p-ethyl was converted into quizalofop acid which was used as hapten and was analyzed by MS, 1H NMR and IR. It was conjugated to bovine serum albumin (BSA) by active ester method (AEM) and ovalbumin (OVA) by mixed anhydride reaction (MAR), respectively. The UV spectra showed that these conjugates were obtained successfully, and the coupling ratio of Hatpen-BSA is 29.23 and the coupling ratio of Hatpen-OVA is 7.87.2. The special polyclonal antibodies against quizalofop-p-ethyl were generated by immunizing 3 New Zealand white rabbits with the hapten-protein conjugate (Hapten-BSA). Titers of 3 rabbits are M4736 (1:80,000), M4740 (1:128,000), M4743 (1:800,000). The titer of M4743 was much higher than those of others, so it was used in the experiment.The two dimensional titration of the antisera M4743 against Hapten-OVA and the ELISA method revealed that the best work concentration is that the dilution of M4743 antiserum was 1/200,000 against 2.05 μg/mL Hapten-OVA, this combinations was used to perform the following ELISA method. The effect of various factors, such as coating medium, the type and concentration of organic solvents, ionic strength, which may influence the afflinity between antibodies and coating antigen and the sensitivity of the assay were studied to optimize analytical conditions of ELISA. With optimized combination, we used the antiserum M4743 to establish IC-ELISA method, a standard curve for quizalofop-p-ethyl was obtained by plotting inhibition rate versus LogC (C is the concentration of quizalofop-p-ethyl). The inhibition rate curve equation is I=31.793LogC+96.314, r=0.994. The assay showed that the IC50 for quizalofop-p-ethyl was 0.03495 μg/mL and the lowest detection limit was 0.00192 μg/mL, the working range was assigned to concentrations 0.002-0.5 ug/mL for quizalofop-p-ethyl. Furthermore, other structurally closely related compounds tested did not...