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目的 为防治白内障术后因晶状体上皮细胞增殖而产生的后发性白内障 ,制备抗家兔晶状体上皮细胞的单克隆抗体 ,从而更进一步以其为载体与细胞毒素偶联 ,特异性抑制晶状体上皮细胞生长而不损伤眼内其他组织 ,为防治后发性白内障奠定实验基础。方法 应用家兔晶状体上皮细胞与佐剂混合 ,免疫 BAL B/ c小鼠 ,通过聚乙二醇 (PEG -40 0 0 )使被免疫的小鼠脾细胞与同系小鼠骨髓瘤细胞融合 ,细胞融合之后 ,经过 HAT选择性培养液培养 ,用间接免疫荧光抗体法和免疫组化 SP法检测杂交瘤细胞上清夜中的抗体 ,筛选出呈阳性反应的杂交瘤细胞 ,再经 3次甲基纤维素克隆化培养以保证单克隆抗体的特性 ,最后将此单克隆抗体对人眼组织进行交叉反应。结果 被免疫的小鼠脾细胞与骨髓瘤细胞 SP2 /0通过 PEG-40 0 0融合后 ,经 HAT选择性培养 ,16 d后杂交瘤细胞克隆产生 ,对其进行 3次甲基纤维素培养基克隆化培养 ,获得检测抗体阳性率达 10 0 %的克隆系为一株。用免疫组化 SP法检测此抗体仅对兔晶状体上皮细胞抗原呈阳性反应 ,而对兔眼角膜、虹膜、人眼角膜、虹膜、晶状体上皮细胞呈阴性反应。结论 成功制备了抗家兔晶状体上皮细胞的单克隆抗体 ,此单克隆抗体具有较强的特异性 ,对人眼组织无交叉反应 ,为进一步导向治疗后发性
Objective To prevent and treat cataract due to the proliferation of lens epithelial cells and the generation of posterior cataract, preparation of anti-rabbit lens epithelial cells monoclonal antibody, which further as its carrier and cytotoxic conjugate, specifically inhibiting lens epithelial cells Growth without damage to other tissues in the eye, to lay the experimental foundation for the prevention and treatment of post-cataract. Methods BALB / c mice were immunized with rabbit lens epithelial cells and adjuvant. The splenocytes of immunized mice were fused with myeloma cells by polyethylene glycol (PEG - 40 000), and the cells After fusion, the cells were cultured in HAT selective culture medium, and the antibodies in the supernatant of the hybridoma cells were detected by indirect immunofluorescence antibody method and immunohistochemical SP method. The hybridoma cells that screened positive were screened, The clone was cultured to ensure the characteristics of the monoclonal antibody, and finally the monoclonal antibody was cross-reacted with the human eye tissue. Results The immunized mouse spleen cells were fused with myeloma cells SP2 / 0 by PEG-400 and selectively cultured by HAT. After 16 days, the hybridoma cells were cloned and subjected to 3 times of methylcellulose medium Clone culture to obtain a detection antibody positive rate of 10 0% of the clone system as a strain. Immunohistochemical SP method detected this antibody only positive for rabbit lens epithelial cells antigen, and rabbit cornea, iris, cornea, iris, lens epithelial cells was negative. Conclusion Monoclonal antibodies against rabbit lens epithelial cells have been successfully prepared. The monoclonal antibodies have strong specificity and no cross-reaction to human eye tissue. In order to further guide the treatment of post-traumatic