Flow Cytometric Analysis of Electronic Nuclear Volume and DNA Content in Normal Mouse Tissues
Flow Cytometric Analysis of Electronic Nuclear Volume and DNA Content in Normal Mouse Tissues
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DOI：
10.4161/cc.3.3.754
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发表时间：
2004-03
期刊：
影响因子：
4.3
通讯作者：
A. Krishan;R. Cabana
中科院分区：
文献类型：
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作者：
A. Krishan;R. Cabana
在流式细胞术中使用光散射以基于细胞的大小和/或粒度来鉴定细胞。然而，前向光散射不是细胞大小的准确量度。利用库尔特原理测量电子体积更准确。然而，EV不能在大多数市售流式细胞仪上测量。本文介绍了一种能同时测量电子核体积（ENV）和DNA含量的流式细胞仪的研制和应用。在本研究中，我们使用市售的NPE QuantaTm测量不同正常小鼠组织的EV和DNA含量。新鲜/冷冻或福尔马林固定石蜡包埋的小鼠组织经处理以分离细胞核，然后分析EV与DNA含量。通过使用这两个参数，在肝脏、胸腺、小肠和骨髓中鉴定出不同的亚群。EV与DNA含量的双参数分析可以是用于鉴定异质细胞混合物中的亚群的有价值的技术，例如复杂组织如骨髓、肠和肿瘤的亚群。所建立的方法是快速的，并可以提供有价值的数据的鉴定和表征亚群的细胞周期分析的流式细胞术。
Light scatter is used in flow cytometry for identification of cells based on their size and/or granularity. However, forward light scatter is not an accurate measure of cell size. The measurement of Electronic Volume (EV) by Coulter principle is more accurate. However, EV cannot be measured on most of the commercially available flow cytometers. We have described the development and applications of a flow cytometer that can simultaneously measure Electronic Nuclear Volume (ENV) and DNA content. In the present study we have used a commercially available NPE QuantaTm for measuring EV and DNA content of different normal mice tissues.Fresh/frozen or formalin fixed-paraffin embedded tissues from mice were processed for isolation of nuclei, which were then analyzed for EV versus DNA content. By using these two parameters, distinct sub-populations were identified in liver, thymus, small intestine and bone marrow. Dual parametric analysis of EV versus DNA content can be a valuable technique for identification of sub-populations in heterogeneous cell mixtures such as those of complex tissues like bone marrow, intestine and tumors. The methods established are rapid and can provide valuable data for identification and characterization of sub-populations for cell cycle analysis by flow cytometry.