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摘要：通过用特异性抗氯噻啉的单克隆抗体淘选噬菌体展示肽库获得优尔种噬菌体展示肽。选择克隆L4-29建立噬菌体酶免疫吸附分析（P-ELISA）和噬菌体化学发光酶免疫分析（P-CLEIA）。 P-ELISA和P-CLEIA的最佳缓冲液为pH 8，0.14 M NaCl和2.5％甲醇的PBS缓冲液。P-ELISA的半数最大抑制浓度（IC50）值，检测限（LOD，IC10值）和线性范围（IC10-IC90）分别为1.45,0.55和0.55至3.82 ng mL -1，P-CLEIA 分别为0.86,0.13和0.13-5.84 ng mL -1。与传统ELISA相比，P-ELISA和P-CLEIA的灵敏度分别提高了40倍和68倍。P-CLEIA比P-ELISA更敏感，具有更宽的线性范围。两种分析方法与氯噻啉的类似物（除吡虫啉）没有显著的交叉反应性。P-ELISA和P-CLEIA两种方法在稻田水，土壤，甘蓝，水稻，苹果，梨和番茄样品中对氯噻啉的添加回收率分别为72.3-101.3％和73.9-102.6％。P-ELISA和P-CLEIA与高效液相色谱（HPLC）对样品中氯噻啉检出结果的相关性良好，可用于实际样品中的氯噻啉的检测。34621
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毕业论文关键词：氯噻啉;酶免疫吸附分析;化学发光酶免疫分析;噬菌体展示肽;模拟表位
Development of phage enzyme immunoassays for the determination of imidaclothiz
Abstract：Six phage-displayed peptides were obtained by biopanning the phage-displayed peptide libraries with the specific anti-imidaclothiz monoclonal antibody. A phage enzyme-linked immunosorbent assay (P-ELISA) and a phage chemiluminescent enzyme immunoassay (P-CLEIA) for imidaclothiz were developed by using a clone L4-29. The optimal buffers of P-ELISA and P-CLEIA were pH 8, PBS buffer containing 0.14 M NaCl and 2.5% methanol. The half maximal inhibition concentration (IC50) values, limit of detection (LOD, IC10 values) and linear range (IC10 to IC90) of the P-ELISA were 1.45, 0.55 and 0.55 to 3.82 ng mL-1, while the P-CLEIA were 0.86, 0.13 and 0.13 to 5.84 ng mL-1, respectively. Compared with conventional ELISAs, the sensitivities of P-ELISA and P-CLEIA were increased by more than 40-fold and 68-fold, respectively. The P-CLEIA was more sensitive and showed wider linear range than the P-ELISA. Two analyses had no significant cross-reactivity with the analogues of imidaclothiz except for imidacloprid. Recoveries of the P-ELISA and P-CLEIA for imidaclothiz in paddy water, soil, cabbage, rice, apple, pakchoi, pear and tomato samples were 72.3-101.3% and 73.9-102.6%, respectively. The P-ELISA and P-CLEIA were feasible to detect the amounts of imidaclothiz in the authentic samples, which showed good correlation with that obtained from high-performance liquid chromatography (HPLC).
Keywords：Imidaclothiz, Enzyme-linked immunosorbent assay, Chemiluminescent enzyme immunoassay, Phage-displayed peptide, Peptidomimetics
目录
摘要 2
关键词 2
Abstract 2
Keywords 2
引言 2
1.材料与方法 4
1.1材料 4
1.2主要仪器 4
1.3方法 4
1.3.1氯噻啉单克隆抗体的制备 4
1.3.2生物淘选 4
1.3.3噬菌体酶免疫吸附分析方法 5
1.3.4噬菌体化学发光酶免疫分析方法 5
1.3.5P-ELISA和P-CLEIA的优化 5
1.3.6标准曲线的建立 5
1.3.7交叉反应性（CR） 5
1.3.8 加标样品的分析 5
1.3.9 HPLC分析验证 6
2.结果与分析 6
2.1噬菌体展示肽的淘选 6
2.2噬菌体展示肽的选择 6
2.3 P-ELISA和P-CLEIA的优化 7
2.4 P-ELISA和P-CLEIA的灵敏度 7
2.5 P-ELISA和P-CLEIA的特异性 8
2.6加标样品的分析 9
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