A simple approach for rapid and cost-effective quantification of extracellular vesicles using a fluorescence polarization technique
A simple approach for rapid and cost-effective quantification of extracellular vesicles using a fluorescence polarization technique
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DOI：
10.1186/s13036-019-0160-9
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发表时间：
2019-04-16
影响因子：
5.6
通讯作者：
Park, Ki Soo
中科院分区：
文献类型：
--
作者：
Kalimuthu, Kalishwaralal;Kwon, Woo Young;Park, Ki Soo
细胞外囊泡（EVs）是由所有细胞积极分泌的磷脂膜结合囊泡。由于它们携带由亲本细胞表达的特定标记物，因此可以通过液体活检来识别特定细胞。为了促进基于ev的临床诊断，一种快速可靠的ev计数方法至关重要。基于亲脂荧光素探针5-十二羧基氨基荧光素（C12-FAM）的荧光偏振（FP）检测，建立了一种快速、经济高效的ev定量方法。C12-FAM的烷基尾部被特异性地整合到电动汽车中，由于缓慢的扩散运动而产生高FP值。我们使用新策略量化了HT29和TCMK1两种细胞系衍生的电动汽车，具有与商业方法相当的良好灵敏度。新方法涉及到最小的复杂性和实际操作时间。此外，FP信号本质上是比率的，并且对环境噪声具有鲁棒性。
Extracellular vesicles (EVs) are membrane-bound phospholipid vesicles actively secreted by all cells. As they carry specific markers expressed by their parental cells, EVs are utilized to identify specific cells via liquid biopsy. To facilitate EV-based clinical diagnosis, a fast and reliable method to count EVs is critical. We developed a method for rapid and cost-effective quantification of EVs which relies on the fluorescence polarization (FP) detection of lipophilic fluorescein probe, 5-dodecanoylamino fluorescein (C12-FAM). The alkyl tail of C12-FAM is specifically incorporated into the EVs, producing high FP values due to a slow diffusional motion. We quantified EVs derived from two cell lines, HT29 and TCMK1 using the new strategy, with good sensitivity that was at par with the commercial method. The new method involves minimal complexity and hands-on time. In addition, FP signaling is inherently ratiometric and is robust against environmental noise.