Duda Dan G, Cohen Kenneth S, Scadden David T, Jain Rakesh K
Steele Laboratory for Tumor Biology, Department of Radiation Oncology, Massachusetts General Hospital, Boston, Massachusetts 02114, USA.
Nat Protoc. 2007;2(4):805-10. doi: 10.1038/nprot.2007.111.
Blood circulating endothelial cells (CECs) and circulating hematopoietic progenitor cells (CPCs) represent two cell populations that are thought to play important roles in tissue vascularization. CECs and CPCs are currently studied as surrogate markers in patients for more than a dozen pathologies, including heart disease and cancer. However, data interpretation has often been difficult because of multiple definitions, methods and protocols used to evaluate and count these cells by different laboratories. Here, we propose a cytometry protocol for phenotypic identification and enumeration of CECs and CPCs in human blood using four surface markers: CD31, CD34, CD133 and CD45. This method allows further phenotypic analyses to explore the biology of these cells. In addition, it offers a platform for longitudinal studies of these cells in patients with different pathologies. The protocol is relatively simple, inexpensive and can be adapted for multiple flow cytometer types or software. The procedure should take 2-2.5 h, and is expected to detect 0.1-6.0% viable CECs and 0.01-0.20% CPCs within blood mononuclear cell population.
循环内皮细胞（CECs）和循环造血祖细胞（CPCs）代表了两个被认为在组织血管化过程中发挥重要作用的细胞群体。目前，CECs和CPCs作为替代标志物，在包括心脏病和癌症在内的十几种病症患者中得到研究。然而，由于不同实验室用于评估和计数这些细胞的多种定义、方法和方案，数据解释往往很困难。在此，我们提出一种细胞计数方案，用于使用四种表面标志物（CD31、CD34、CD133和CD45）对人血液中的CECs和CPCs进行表型鉴定和计数。该方法允许进行进一步的表型分析，以探索这些细胞的生物学特性。此外，它为不同病症患者中这些细胞的纵向研究提供了一个平台。该方案相对简单、成本低廉，并且可以适用于多种类型的流式细胞仪或软件。该程序应耗时2 - 2.5小时，预计可在血液单核细胞群体中检测到0.1 - 6.0%的存活CECs和0.01 - 0.20%的CPCs。
