An accurate, precise method for general labeling of extracellular vesicles.
An accurate, precise method for general labeling of extracellular vesicles.
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DOI：
10.1016/j.mex.2015.08.002
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发表时间：
2015
期刊：
影响因子：
1.9
通讯作者：
Searles CD
中科院分区：
文献类型：
--
作者：
Gray WD;Mitchell AJ;Searles CD
钙黄素AM是非荧光的，直到它被动地进入EVS，之后它被激活，变成荧光的和EV不意味着的。穿透性/裂解EVS与抗体和亲脂膜染色呈阳性，而钙黄绿素未见标记。与使用抗体或膜染色的方法不同，钙黄素AM允许区分完整的EVS和碎片。钙黄素AM可用于从多种细胞类型中检测完整的EVS。细胞外，膜泡(微囊泡、外切体)由细胞分泌，可作为细胞间通讯的媒介。用流式细胞术检测它们的方法包括使用荧光染色水泡膜的试剂，或针对特定细胞起源抗原的荧光抗体。然而，这些方法可能会错误地检测细胞碎片或需要事先了解细胞起源。在这里，我们证明了钙黄绿素AM适合于通过流式细胞术检测完整的细胞外小泡(EVS)。
Calcein AM is non-fluorescent until it passively enters EVs, after which it is activated and becomes fluorescent and EV-impermeant. Permeabilized/lysed EVs label positive with antibodies and lipophilic membrane stain, whereas no labeling was observed with calcein. In contrast to methods that use antibodies or membrane stains, calcein AM allows for the differentiation between intact EVs and debris. Calcein AM can be used for detection of intact EVs from numerous cell types. Extracellular, membrane vesicles (microvesicles, exosomes) are secreted by cells and may serve as mediators of intercellular communication. Methods for detecting them by flow cytometry have included the use of agents that fluorescently stain vesicle membrane, or fluorescent antibodies that target specific cell-of-origin antigens. However, these methods may falsely detect cell debris or require prior cell-of-origin knowledge. Here, we demonstrate the suitability of calcein AM for detection of intact extracellular vesicles (EVs) by flow cytometry.