兔抗人DR5抗血清诱导Jurkat细胞凋亡

制备兔抗人sDR5抗血清,检测它对Jurkat细胞的生长抑制和凋亡诱导作用。采用本室制备的sDR5免疫新西兰白兔,制备兔抗人sDR5抗血清,用ELISA法测定抗sDR5抗血清效价及抗血清的特异性。MTT试验分析它对Jurkat细胞生长抑制影响,倒置光显微镜和荧光显微镜观察抗sDR5抗血清对Jurkat细胞形态的影响,用AnnexinV/PI双染试剂盒检测Jurkat细胞凋亡率,琼脂糖凝胶电泳检测Jurkat细胞中DNA的片断化。结果:获得了高效价特异性兔抗人sDR5抗血清。兔抗人sDR5抗血清对Jurkat细胞具有显著的细胞生长抑制作用,并呈剂量依赖性。兔抗人sDR5抗血清处理后,Jurkat细胞可出现典型的细胞凋亡的形态特征:细胞膜皱缩,出泡,染色质浓缩,形成凋亡小体等。流式细胞术结果显示:兔抗人sDR5血清1/80、1/160作用Jurkat细胞2 h,细胞凋亡率分别为54.98%和34.13%。兔抗人sDR5抗血清可导致Jurkat细胞中的DNA片段化。本室制备的兔抗人sDR5抗血清能抑制Jurkat细胞生长和诱导Jurkat细胞凋亡。

The apoptosis of Jurkat cells induced by rabbit antiserum against human sDR5

To prepare the rabbit antiserum against human sDR5 and to observe its effect on growth inhibition and apoptosis of Jurkat cells.The purified human sDR5 was used as antigen to immunize New Zealand white rabbits in order to prepare the specific anti-human sDR5 antibody.The specificity of this antibody was analyzed by ELISA and its effect on the morphology of Jurkat cells was observed by light and fluorescent microscopies.Meanwhile,the effect on the growth inhibition of Jurkat cells was examined by MTT assay,and the percentage of apoptosis in Jurkat cells was determined by flow cytometry using the annexin V/PI double staining kit.The DNA fragmentation in Jurkat cells was analyzed by agarose gel electrophoresis.In this way,specific rabbit antiserum against human sDR5 with high titer(1∶100 000) could be obtained.The excellent growth inhibition effect of this antibody on Jurkat cells could be demonstrated in a dose-dependent manner.In addition,the Jurkat cells after treatment with rabbit anti-human sDR5 antiserum showed the typical morphological characteristics of apoptosis,including membrane crenation,bubbling,chromatin condensation and formation of apoptotic bodies.As demonstrated by flow cytometry,the apoptotic rates of Jurkat cells after treatment with 1/80 and 1/160 of rabbit anti-human sDR5 sera for 2 hrs were 54.98% and 34.13% respectively.It was also evident from agarose gel electrophoresis that the DNA fragmentation in Jurkat cells occurred after treatment with this rabbit antiserum.From this observation,it is clear that the rabbit anti-human sDR5 serum efficiently induce growth inhibition and apoptosis of Jurkat cells.