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Production and Identification of Broad Spectrum Monoclonal Antibody against a Group of Pyrethroid Insecticides / 分析化学
Chinese Journal of Analytical Chemistry ; (12): 1245-1251, 2014.
Article in Chinese | WPRIM | ID: wpr-456490
ABSTRACT
The objective of this study is to generate broad spectrum monoclonal antibody ( mAb ) against a group of pyrethroid insecticides and to identify its immunological characteristics. The generic hapten 3-phenoxy-benzoic acid ( PBA) was conjugated to carrier protein BSA by activated ester method. Balb/c mice were immunized with PBA-BSA. The titer of polyclonal antibody ( pAb ) was detected by indirect enzyme-linked immunosorbent assay ( ELISA) after five times immunization. The mouse with high titer and sensitivity was selected for cell fusing. The splenocytes of immunized mice were fused with Sp2/0 cells and the cultural supernatants of hybridoma cells were screened by indirect non-competitive ELISA based on the coating antigen PBA-ovoalbumin ( PBA-OVA ) . High-sensitivity and high-specificity mAb was prepared after subcloning using limiting dilution method. Purified mAb was obtained after purified by saturated ammonia sulfate precipitation and protein G affinity column. The immunological characteristics of mAb such as titer, antibody subtypes, affinity constant and the sensitivity to pyrethroid insecticides were characterized by indirect ELISA; The results of UV spectroscopy and SDS-PAGE showed that PBA-BSA artificial antigen was synthesized successfully. A hybridoma cell line (4H11) secreting anti-pyrethroid mAb was established. The titre of ascites was up to 16. 5×106, and the mAb was IgG1 subtype. The affinity constant of the mAb to PBA was about 2. 5×107 L/mol, with a IC50 value of 208. 83 μg/L and a detection limit of 21. 23 μg/L to PBA. Simultaneously, beta-cypermethrin, flucythrinate, cypermethrin and fenvalerate were sensitively recognized by the mAb with the IC50 of 1. 01, 2. 15, 3. 16 and 3. 67μg/L, respectively.

Full text: Available Index: WPRIM (Western Pacific) Type of study: Diagnostic study / Prognostic study Language: Chinese Journal: Chinese Journal of Analytical Chemistry Year: 2014 Type: Article

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Full text: Available Index: WPRIM (Western Pacific) Type of study: Diagnostic study / Prognostic study Language: Chinese Journal: Chinese Journal of Analytical Chemistry Year: 2014 Type: Article