The anti-chlorpyrifos polyclonal antibodies were obtained by using the artificial immuneantigen to immune in New Zealands white rabbits. The enzyme-tagged antibodies wereprepared by coupling horseradish peroxidase (HR...The anti-chlorpyrifos polyclonal antibodies were obtained by using the artificial immuneantigen to immune in New Zealands white rabbits. The enzyme-tagged antibodies wereprepared by coupling horseradish peroxidase (HRP) to the purified antibody with themodified sodium periodate method. The indirect competitive enzyme linked immuno-sorbentassays (ELISA) and the HRP-tagged antibody direct ELISA (E-Ab) were established, respectively.The limit of detection (LOD) for the indirect ELISA and E-Ab were 0.0033 and 0.0042 gmL-1, respectively. The linear detection ranged well from 0.005 to 2.0 g mL-1.展开更多
To produce high-affinity monoclonal antibodies against pesticide carbofuran, and the develop immunochemical assays for people＇s health and environmental protection, the hapten 4-[[（2,3-dihydro-2,2-dimethyl-7-benzofu...To produce high-affinity monoclonal antibodies against pesticide carbofuran, and the develop immunochemical assays for people＇s health and environmental protection, the hapten 4-[[（2,3-dihydro-2,2-dimethyl-7-benzofuranyloxy） carbonyl]- amino]-butanoic acid （BFNB） of carbofuran was synthesized and Balb/c mice were immunized by the hapten-carrier （BFNB-bovine serum albumin, BFNB-BSA） conjugates. The splenocytes of immunized mice were fused with Sp2/0 cells and the cultural supernatants of hybridoma cells were screened by the indirect enzyme-linked immunoabsorbent assay （ELISA）, based on BFNB-ovoalbumin conjugates （BFNB-OVA）. Purified monoclonal antibody （McAb） was obtained from fluids of ascites, deposited by octanoic acid and ammonium sulfate. The affinity and the specificity of McAb were characterized by ELISA or indirect competitive ELISA. A hybridoma cell line （5D3） secreting anti-carbofuran McAb had been established. The titer of culture medium and ascites was up to 1：2.048 × 10^3 and 1：1.024 × 10^6, respectively, and the subtype of the McAb was IgG1. The affinity constant of the McAb was about 2.54 × 10×9 L mol^-1, with an IC50 value of 1.18 ng mL^-1 and a detection limit of 0.01 ng mL^-1. Cross-reactivity studies showed that the McAb was quiet specific for carbofuran, as among the four analogous compounds, they were all hardly recognized （4.59 × 10^-4% for 2,3-dihydro-2,2- dimethyl-7-benzofuranol and less than 3.0 × 10^4% for others）. The prepared McAb had a very high affinity and specificity, and it could be used to develop ELISA for rapid determination of carbofuran.展开更多
文摘The anti-chlorpyrifos polyclonal antibodies were obtained by using the artificial immuneantigen to immune in New Zealands white rabbits. The enzyme-tagged antibodies wereprepared by coupling horseradish peroxidase (HRP) to the purified antibody with themodified sodium periodate method. The indirect competitive enzyme linked immuno-sorbentassays (ELISA) and the HRP-tagged antibody direct ELISA (E-Ab) were established, respectively.The limit of detection (LOD) for the indirect ELISA and E-Ab were 0.0033 and 0.0042 gmL-1, respectively. The linear detection ranged well from 0.005 to 2.0 g mL-1.
基金This study was supported by the Key Project of Natural Science Foundation of Guangdong Province, China （036842） Key Project of Key Technologies R＆D Program of Guangzhou City, China （2004Z2-E0031 ,2004Z1-E0061）+2 种基金 Natural Science Foundation of Guangdong Province, China （06300421） National HighTech Research and Development Program of China （863 Program） （2006AA10Z447） and Key Technologies R＆D Program during the 10th Five-Year Plan Period, China （2002BA518A06-04）.
文摘To produce high-affinity monoclonal antibodies against pesticide carbofuran, and the develop immunochemical assays for people＇s health and environmental protection, the hapten 4-[[（2,3-dihydro-2,2-dimethyl-7-benzofuranyloxy） carbonyl]- amino]-butanoic acid （BFNB） of carbofuran was synthesized and Balb/c mice were immunized by the hapten-carrier （BFNB-bovine serum albumin, BFNB-BSA） conjugates. The splenocytes of immunized mice were fused with Sp2/0 cells and the cultural supernatants of hybridoma cells were screened by the indirect enzyme-linked immunoabsorbent assay （ELISA）, based on BFNB-ovoalbumin conjugates （BFNB-OVA）. Purified monoclonal antibody （McAb） was obtained from fluids of ascites, deposited by octanoic acid and ammonium sulfate. The affinity and the specificity of McAb were characterized by ELISA or indirect competitive ELISA. A hybridoma cell line （5D3） secreting anti-carbofuran McAb had been established. The titer of culture medium and ascites was up to 1：2.048 × 10^3 and 1：1.024 × 10^6, respectively, and the subtype of the McAb was IgG1. The affinity constant of the McAb was about 2.54 × 10×9 L mol^-1, with an IC50 value of 1.18 ng mL^-1 and a detection limit of 0.01 ng mL^-1. Cross-reactivity studies showed that the McAb was quiet specific for carbofuran, as among the four analogous compounds, they were all hardly recognized （4.59 × 10^-4% for 2,3-dihydro-2,2- dimethyl-7-benzofuranol and less than 3.0 × 10^4% for others）. The prepared McAb had a very high affinity and specificity, and it could be used to develop ELISA for rapid determination of carbofuran.
