本文選題：-戊基苯酚(-AP) + 單克隆抗體�。� 參考：《細(xì)胞與分子免疫學(xué)雜志》2017年03期

【摘要】：目的制備4-戊基苯酚(4-AP)單克隆抗體(mAb),通過(guò)分子生物學(xué)方法對(duì)其可變區(qū)cDNA克隆和同源建模,為4-AP單鏈抗體制備及抗體分子層面改造奠定基礎(chǔ),同時(shí)為4-AP的免疫學(xué)檢測(cè)提供一種新方法。方法將免疫小鼠脾臟細(xì)胞與Sp2/0細(xì)胞融合,篩選出F10陽(yáng)性單抗細(xì)胞株。通過(guò)其mRNA的抽提,克隆出可變區(qū)cDNA序列,利用生物軟件進(jìn)行同源建模和分子對(duì)接。結(jié)果在最適條件下,其間接競(jìng)爭(zhēng)ELISA(ic-ELISA)公式為y=A_2+(A_1-A_2)/(1+(x/x_0)~p)(A_1=1.28;A_2=-0.07;x_0=12 560.75;p=0.74),相關(guān)系數(shù)(R~2)為0.997,其最低檢測(cè)限為0.65μg/mL。mAb重鏈和輕鏈分別屬于IgG1型和Kappa型,其與4-AP的結(jié)合力為氫鍵和疏水作用。結(jié)論成功制備出1株穩(wěn)定分泌的4-AP mAb的雜交瘤細(xì)胞株。通過(guò)同源建模對(duì)抗體可變區(qū)空間結(jié)構(gòu)進(jìn)行深入研究,為單鏈抗體及抗體改造提供參考,同時(shí)為4-AP含量檢測(cè)提供新方法。
[Abstract]:Objective to prepare the monoclonal antibody of 4-pentylphenol 4-APs and clone its variable region cDNA by molecular biology, and to establish the foundation for the preparation of 4-AP single chain antibody and the modification of its molecular level. It also provides a new method for immunological detection of 4-AP. Methods spleen cells of immunized mice were fused with Sp2/0 cells and F 10 positive McAb cell lines were screened. Through the extraction of its mRNA, the variable region cDNA sequence was cloned, and the homologous modeling and molecular docking were carried out by using biological software. Results under the optimum conditions, the indirect competition ELISA-ELISA-ELISA-ELISAA) formula is y=A_2 / A1-A / 2 / T / 1 / T / 1 / X / 0 / T / 1 / 1 / 0 / 0 / 1 / 1 / 1 / 1 / 0 / 1 / 1 / 1 / 1 / 1 / 1 / 1 / 0 / 0 / 0 / 0 / 0 / 12 560.75p0.74. the minimum detection limit is 0.997. the minimum detection limit is 0.65 渭 g/mL.mAb heavy chain and Kappa type, respectively, and its binding force with 4-AP is hydrogen bond and hydrophobic interaction. The results show that the maximum detection limit is 0.65 渭 g/mL.mAb heavy chain and Kappa type respectively, and its binding force with 4-AP is hydrogen bond and hydrophobicity. Conclusion A hybridoma cell line with stable secretion of 4-AP mAb was successfully prepared. The spatial structure of variable region of antibody was studied by homology modeling, which provided a reference for scFV and antibody modification, and a new method for 4-AP content detection.
【作者單位】： 華東理工大學(xué)生物反應(yīng)器工程國(guó)家重點(diǎn)實(shí)驗(yàn)室;上海出入境檢測(cè)檢驗(yàn)局工業(yè)產(chǎn)品和原材料檢測(cè)中心;
【基金】：上海檢驗(yàn)檢疫局科技計(jì)劃(Hk016-2014)
【分類(lèi)號(hào)】：R392

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