高有机溶剂耐受性的苯并(a)芘单克隆抗体制备及其免疫反应特性研究

目的 制备适应油脂样品中苯并(a)芘Benzo(a)pyrene,BaP]检测需求的抗BaP单克隆抗体。方法 利用杂交瘤单克隆抗体技术制备抗BaP的抗体,在抗体筛选过程中使用高浓度的有机溶剂(50%甲醇)来选择具有较高有机溶剂耐受能力的抗体,并利用间接竞争酶联免疫吸附法考察了抗体的免疫反应特性。结果 筛选获得一株对有机溶剂有较高耐受能力的、稳定分泌BaP单克隆抗体的细胞株2H10,该抗体为IgG1亚型,可耐受高浓度的甲醇、丙酮、二甲亚砜(Dimethyl sulfoxide,DMSO)和二甲基甲酰胺(Dimethylformamide,DMF)。60%甲醇和60% DMSO作为样品稀释液时,抗体对BaP的IC₅₀分别为81.4和24.5 ng/mL,以60% DMSO作为样品稀释液,可以显著降低BaP抗体对其它多环芳烃化合物的交叉反应率。结论 本研究获得高特异性的BaP人工抗原及高有机溶剂耐受性的抗BaP单克隆抗体。

Preparation of monoclonal antibody to benzo(a)pyrene with high organic solvent tolerance and characterization of its immune response

Objective To prepare monoclonal antibodies against benzo(a)pyrene (BaP) to meet the needs of BaP detection in oil samples. Methods A monoclonal antibody against BaP were prepared by hybridoma technique. A high concentration of organic solvent (50% methanol) was used in the antibody screening process to select antibodies with high tolerance to organic solvents, and the immunoreactivity characteristics of the antibody were investigated by indirect competitive enzyme-linked immunosorbent assay. Results A cell line 2H10 with high tolerance to organic solvents and stable secretion of monoclonal antibody against BaP was screened. The antibody (IgG1 isotype) could tolerate high concentrations of methanol, acetone, dimethyl sulfoxide (DMSO), and dimethylformamide (DMF). The half maximal inhibitory concentrations (IC50) of the antibody against BaP were 81.4 and 24.5 ng/mL when 60% methanol and 60% DMSO were used as the sample diluents, respectively. Using 60% DMSO as the sample diluent reduced the cross-reactivities of the BaP antibody against other polycyclic aromatic hydrocarbons compounds. Conclusion In this study, an artificial antigen with high specificity of BaP and an anti-BaP monoclonal antibody with high tolerance to organic solvents have been obtained.