Development of a novel flow cytometric cell-mediated cytotoxicity assay using the fluorophores PKH-26 and TO-PRO-3 iodide
Development of a novel flow cytometric cell-mediated cytotoxicity assay using the fluorophores PKH-26 and TO-PRO-3 iodide
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DOI：
10.1016/s0022-1759(01)00336-2
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发表时间：
2001-06-01
影响因子：
2.2
通讯作者：
Wilkinson, RW
中科院分区：
文献类型：
--
作者：
Lee-MacAry, AE;Ross, EL;Wilkinson, RW
已开发了流式细胞术（FCM）测定细胞介导的细胞毒性（CMC）。在测定中，在与脾细胞效应细胞孵育之前，用膜染料PKH-26标记靶肿瘤细胞群。通过加入活力探针TO-PRO-3碘化物（TP 3）评估靶群体中的细胞死亡，并通过流式细胞术进行分析。细胞毒性的程度通过仅用PKH-26标记的活靶细胞和用PKH-26和TP 3两者标记的死透化细胞的相对数量来确定。这种CMC方法允许在单细胞基础上进行分析，并克服了对放射性化学物质的需求。该通信表明，FCR;I检测试剂盒是一种准确且可重现的实验系统，能够分析自然杀伤（NK）细胞和抗体依赖性细胞介导的细胞毒性。该方法与铬释放试验相当。我们相信这是基于FCM的抗体依赖性细胞介导的细胞毒性（ADCC）测定的首次演示之一，（C）2001 Elsevier Science B. V.保留所有权利。
A flow cytometric (FCM) assay has been developed for the determination of cell-mediated cytotoxicity (CMC). In the assay, the target tumour cell population was labelled with a membrane dye, PKH-26, prior to incubation with splenocyte effector cells. Cell death within the target population was assessed by the addition of the viability probe TO-PRO-3 iodide (TP3) and analysed by flow cytometry. The extent of cytotoxicity was determined by the relative number of live target cells labelled with PKH-26 only and dead, permeabilised cells labelled with both PKH-26 and TP3. This CMC method allows the analysis to be conducted on a single cell basis and overcomes the need for radiochemicals. This communication indicates that the FCR;I assay is an accurate and reproducible experimental system capable of analysing natural killer (NK) cell and antibody-dependent cell-mediated cytotoxicity. The procedure is comparable to the chromium release assay. We believe that this is one of the first demonstrations of an FCM-based antibody-dependent cell-mediated cytotoxicity (ADCC) assay, (C) 2001 Elsevier Science B.V. All rights reserved.