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目的:探讨肿瘤坏死因子相关凋亡诱导配体(TRAIL)与细胞膜微结构域脂筏的关系.方法:采用间接免疫荧光流式细胞 术,分析K562细胞表面TRAIL分子的表达.用FITC标记的霍乱毒素B亚单位(CTx)对K562细胞的脂筏进行染色;用抗FITC 单克隆抗体(mAb)交联脂筏后,以兔抗TRAIL分子抗体及Cy3标记的羊抗兔抗体进行间接免疫荧光染色,激光共聚焦显微镜 分析TRAIL分子与脂筏微结构域的关系.结果:FITC CTx和抗FITCmAb可使脂筏发生交联.脂筏交联的同时TRAIL分子发生 聚集.动态观察表明,抗FITC抗体作用20min后,脂筏发生交联,作用30min时交联最明显;随着脂筏的交联,TRAIL分子发 生聚集.抗FITC抗体作用40min后,脂筏交联程度减弱,且TRAIL分子逐渐被排除于脂筏之外.结论:抗FITC抗体可用于 CTx FITC脂筏染色后脂筏交联的研究.TRAIL分子可能与细胞膜脂筏微结构域有关.
Objective: To investigate the relationship between tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and lipid rafts in cell membrane microdomains.Methods: Indirect immunofluorescence flow cytometry was used to analyze the expression of TRAIL on K562 cells.FITC-labeled cholera Toxin B subunit (CTx) was used to stain lipid rafts of K562 cells. After anti-FITC monoclonal antibody (mAb) cross-linked lipid rafts, indirect immunofluorescence staining was performed with rabbit anti-TRAIL antibody and Cy3-labeled goat anti-rabbit antibody , Confocal laser scanning microscopy analysis of the relationship between TRAIL molecules and lipid raft microstructure.Results: FITC CTx and anti-FITCmAb can cross-linked lipid rafts.At the same time lipid-raft cross-linked TRAIL molecules aggregation.Dynamic observation showed that anti-FITC antibody After 20 min, the lipid rafts crosslinked and the most obvious cross-linking was observed at 30 min. With the cross-linking of lipid rafts, the TRAIL molecules aggregated. After 40 min anti-FITC antibody treatment, the degree of cross-linking of lipid rafts was weakened and the TRAIL molecules were gradually Excluding anti-lipid rafts.Conclusion: Anti-FITC antibody can be used to study lipid-raft cross-linking after lipid raft staining by CTx FITC.TRAIL molecules may be related to the lipid raft microstructure of the cell membrane.