Gool Elmar L, Stojanovic Ivan, Schasfoort Richard B M, Sturk Auguste, van Leeuwen Ton G, Nieuwland Rienk, Terstappen Leon W M M, Coumans Frank A W
Department of Biomedical Engineering and Physics;
Department of Clinical Chemistry.
Clin Chem. 2017 Oct;63(10):1633-1641. doi: 10.1373/clinchem.2016.271049. Epub 2017 Aug 7.
Identification, enumeration, and characterization of extracellular vesicles (EVs) are hampered by the small size of EVs, a low refractive index, and low numbers of antigens on their surface.
We investigated the potential of a 48-multiplex surface plasmon resonance imaging (SPRi) system to perform EV phenotyping. Antigen surface density of 11 antigens was measured on the human breast cancer cell lines HS578T, MCF7, and SKBR3 and their EVs by use of both SPRi and the widely used flow cytometry (FCM).
For cells, the SPRi and FCM signals for antigen exposure correlated (R = 0.66, R = 0.78, R = 0.60). With regard to EVs, SPRi detected 31 out of 33 tested antibody-EV pairs, whereas our flow cytometer detected 5 antibody-EV pairs because of high blank and isotype control signals. For HS578T-derived EVs, the SPRi and FCM signals correlated (R = 0.98). However, on MCF7- and SKBR3-derived EVs, insufficient antigens were detected by our flow cytometer. To confirm that the SPRi responses correlated with mean antigen density on EVs, the SPRi responses of EVs were correlated with antigen density on parental cells as measured by FCM (R = 0.77, R = 0.49, R = 0.52).
SPRi responses correlate with mean antigen density. Moreover, SPRi detects lower antigen-exposure levels than FCM because SPRi measures an ensemble of EVs binding to the sensor surface, whereas FCM detects antigens of single EV.
细胞外囊泡（EVs）的鉴定、计数和表征受到其体积小、折射率低以及表面抗原数量少的阻碍。
我们研究了一种48通道表面等离子体共振成像（SPRi）系统用于进行EVs表型分析的潜力。通过使用SPRi和广泛应用的流式细胞术（FCM），测量了人乳腺癌细胞系HS578T、MCF7和SKBR3及其EVs上11种抗原的表面密度。
对于细胞，抗原暴露的SPRi和FCM信号具有相关性（R = 0.66、R = 0.78、R = 0.60）。对于EVs，SPRi在33对测试的抗体-EV对中检测到31对，而我们的流式细胞仪由于高空白和同型对照信号仅检测到5对抗体-EV对。对于HS578T衍生的EVs，SPRi和FCM信号具有相关性（R = 0.98）。然而，对于MCF7和SKBR3衍生的EVs，我们流式细胞仪检测到的抗原不足。为了证实SPRi反应与EVs上的平均抗原密度相关，将EVs的SPRi反应与通过FCM测量的亲代细胞上的抗原密度进行相关性分析（R = 0.77、R = 0.49、R = 0.52）。
SPRi反应与平均抗原密度相关。此外，SPRi检测到的抗原暴露水平低于FCM，因为SPRi测量的是结合到传感器表面的一组EVs，而FCM检测的是单个EV的抗原。
