抗角蛋白单克隆抗体对舌鳞癌细胞角蛋白酪氨酸磷酸化的作用

目的观察抗角蛋白抗体对角蛋白酪氨酸磷酸化的影响。方法以鼠抗人角蛋白的IgG单克隆抗体作用于培养中的Tca8113,以抗乙肝病毒表面抗原抗体做阴性对照,同时设置阳性对照和空白对照。细胞固定、包埋后以胶体金标记的羊抗鼠IgG抗体染色,用透射电镜观察。提取抗体作用过Tca8113细胞角蛋白,以抗乙肝病毒表面抗原抗体作用的细胞做阴性对照。不同分子量的角蛋白经SDSPAGE分离后转膜,用抗酪氨酸磷酸化抗体作用、显色。结果抗角蛋白抗体作用的Tca8133细胞胞质的中间丝有胶体金颗粒,与阳性对照组类似。阴性对照组及空白对照均未见胶体金颗粒。转膜后染色,在分子量大约58KD处出现阳性条带。阴性对照未出现条带。结论抗角蛋白抗体结合于胞浆中间丝,58KD角蛋白发生酪氨酸磷酸化。

Effects of Monoclonal Anti-keratin Antibody on Keratin of Live Human Tca Cell line

Objective To study effects of antikeratin Monoclonal antibody on keratin of live Human Tca cells.Methods Human Tca cells were primarily cultured in vitro with anti-keratin antibodies,and some other Tca cells cultured with monoclonal antibodies to HbsAg served as the negative group.Meanwhile the positive group and the blank group were also established.All the cells were fixed and embedded before stained with immunogold labeled anti-mouse antibodies.Then the cells were observed under electron microscope.Extract keratin from Tca cells cultured with anti-keratin antibodies.Keratin extracted from Tca cells with HbsAgs served as the negative group.Keratin of different molecular were separated by SDS-PAGE and transfered to nitrocellulose membrane(NC).And then keratin on NC dyed after reacted with Phosphotyrosine Ab. Results The immunogold staining appeared on the keratin intermediates of the cells cultured with anti-keratin antibodies.These phenomena were similar to what were seen in the positive group.While immunogold staining were not seen in cells of the nagetive and blank groups keratin of about 58KD on NC appeared after transferring and dying. Conclusion Anti-keratin antibodies located in keratin intermediates of live Tca cells.The tyrosine phosphorylation of 58KD keratin occurs.