McCloskey T W, Oyaizu N, Coronesi M, Pahwa S
Department of Pediatrics, North Shore University Hospital-Cornell University Medical College, Manhasset, New York 11030.
Clin Immunol Immunopathol. 1994 Apr;71(1):14-8. doi: 10.1006/clin.1994.1045.
Apoptosis is an active process of cellular self-destruction which plays an important physiologic role in the maintenance of homeostasis. This form of cell death has classically been assessed by the appearance of a "ladder-like" banding pattern upon gel electrophoresis of disrupted cells. The electrophoretic method is qualitative but offers no means of quantitation. We have optimized a flow cytometric method which allows quantitation of the apoptotic subset of cells. This assay uses propidium iodide and therefore can be performed on any flow cytometer with a blue-green excitation line. We initially verified the results of this method by using human thymocytes treated with dexamethasone, a known inducer of apoptosis. Gels were run simultaneously with flow cytometric analysis and assessed for the presence or the absence of apoptotic cells. Results obtained with gel electrophoresis correlated well with the flow cytometric method. In further studies, we applied this assay to human PBMC and obtained similar results. In conclusion, this assay is a significant improvement over currently available methodology for the determination of apoptosis. We have found it to be quantitative, reproducible, and applicable to different human cell types. The role of apoptosis in human disease remains to be elucidated and this method offers a convenient means to study this process.
细胞凋亡是一种细胞自我毁灭的主动过程，在维持体内平衡中发挥着重要的生理作用。这种细胞死亡形式传统上是通过对破碎细胞进行凝胶电泳时出现的“梯状”条带模式来评估的。电泳方法是定性的，但无法进行定量分析。我们优化了一种流式细胞术方法，该方法可以对细胞凋亡亚群进行定量分析。该检测方法使用碘化丙啶，因此可以在任何具有蓝绿色激发线的流式细胞仪上进行。我们最初通过使用用地塞米松处理的人胸腺细胞（地塞米松是一种已知的细胞凋亡诱导剂）来验证该方法的结果。在进行流式细胞术分析的同时进行凝胶电泳，并评估凋亡细胞的存在与否。凝胶电泳获得的结果与流式细胞术方法相关性良好。在进一步的研究中，我们将该检测方法应用于人类外周血单个核细胞（PBMC），并获得了类似的结果。总之，该检测方法相对于目前可用的细胞凋亡测定方法有显著改进。我们发现它具有定量性、可重复性，并且适用于不同的人类细胞类型。细胞凋亡在人类疾病中的作用仍有待阐明，而这种方法为研究这一过程提供了一种便捷的手段。
