【作者】 牟文凤；

【导师】 王斌；

【作者基本信息】 青岛大学 ， 病原生物学， 2005， 硕士

【摘要】 ①目的:运用基于SELDI(Surface Enhanced Laser Desorption/Ionization)时间飞行质谱仪和配套的蛋白质芯片技术检测体外培养的肝癌细胞株与正常肝细胞株蛋白质的差异表达,从而为进一步研究肝癌的发病机理奠定基础。②方法:常规培养正常肝细胞株(LO2)和肝癌细胞株(HEPG2),在细胞生长状态良好时收获细胞,加入细胞裂解液裂解。裂解上清蛋白定量后应用IMAC3(固定金属亲和)和WCX2(弱阳离子交换)芯片,SELDI-TOF-MS技术检测两种细胞蛋白质组学的差异。③结果:体外培养的肝癌细胞株与正常肝细胞株的蛋白质存在差异表达,共有20个蛋白表达水平发生变化。IMAC3芯片发现差异峰7个,其中7979,8492,13428,15940 Da在肝癌细胞中表达增高,而8103,10160,11304 Da表达降低;WCX2芯片发现差异峰14个,其中7517,7945,7979Da在肝癌细胞中表达增高,5061,5551,5818,8428,10100,10312,11081,11621,11662,11830,12772 Da表达降低。7979Da蛋白峰可同时被IMAC3及WCX2两种芯片铺获,而且11081Da蛋白峰符合钙结合蛋白S100A10,它可能与p36作用而参与肝癌的发生。④结论:SELDI蛋白芯片技术检测肝癌细胞株与正常肝细胞株蛋白质的差异表达方法简便,敏感性高,重复性好,从而为从蛋白质水平研究肝癌的发病机理及新的治疗靶位的寻找奠定了一定的基础。更多还原

【Abstract】 Objective To detect different protein expression in malignant and normal liver cell lines in vitro using the SELDI ProteinChip platform .which could be used as a basis to study the pathogenesis of liver cancer. Methods Two cell lines , human normal liver cell line L02 and hepatoma cell line HepG2 were cultured routinely and harvested in good conditions using cell lysis. After quantification ,the supernatant of the lysate was tested by IMAC3(Immobilized mental affinity capture) and WCX2(weak cation exchange) chip on SELDI-TOF-MS proteinchip reader respectively. Results The protein expression was different between malignant and normal liver cell lines . Total 20 different expressed proteins were found, among which ,7 were captured by IMAC3 chip and 14 by WCX2 chip. Peaks of 7979, 8492, 13428, and 15940 Da were upragulated and 8103, 10160, 11304 Da downregulated in HepG2 cells by IMAC3 chip ; 7517, 7945, 7979Da were upragulated and 5061, 5551, 5818, 8428, 10100, 10312, 11081, 11621, 11662, 11830, 12772 Da downregulated in HepG2 cells by WCX2 chip respectively. Interestingly, the 7979Da peak was captured by both chips . In addition,the 11081 Da peak was corresponded precisely to the molecular mass of calcium binding protein S100A10, which may participate in the formation of liver cancer in company with p36. Conclusion Detecting different protein expression in malignant and normal liver cell lines using the SELDI ProteinChip platform was simple ,sensitive and repeatable; the results we got could be served as a basis to study the pathogenesis and help the discovery of new therapy targets of liver cancer.更多还原