Extracellular vesicle measurements with nanoparticle tracking analysis - An accuracy and repeatability comparison between NanoSight NS300 and ZetaView
Extracellular vesicle measurements with nanoparticle tracking analysis - An accuracy and repeatability comparison between NanoSight NS300 and ZetaView
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DOI：
10.1080/20013078.2019.1596016
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发表时间：
2019-01-01
影响因子：
16
通讯作者：
von Strandmann, Elke Pogge
中科院分区：
文献类型：
--
作者：
Bachurski, Daniel;Schuldner, Maximiliane;von Strandmann, Elke Pogge
细胞外囊泡（EV）研究领域的不断扩大，需要重复性和准确的方法来表征单个EV。纳米颗粒跟踪分析（NTA）通常用于确定EV浓度和直径。由于EV领域缺乏容易确认和验证NTA数据的方法，因此质疑测量的可靠性仍然非常重要。在这方面，尚未进行不同NTA设备（如马尔文的NanoSight NS 300或Particle Meetry的ZetaView）之间的测量质量比较。为了评估这两种设备的尺寸和浓度测定的准确性和可重复性，我们采用了比较方法，包括透射电子显微镜（TEM）和单粒子干涉反射成像传感（SP-IRIS）的ExoView。使用来自人血清和细胞培养上清液的纳米球、脂质体和超浓缩EV进行多次测试测量。此外，还测量了系列稀释和冻融循环依赖性EV降低，以确定各系统的耐用性。引人注目的是，NanoSight NS 300显示出对聚苯乙烯和二氧化硅纳米球浓度的2.0-2.1倍高估。通过测量EV样品的系列稀释液，我们证明了ZetaView（% BIAS范围：2.7-8.5）与NanoSight NS 300（% BIAS范围：32.9-36.8）相比浓度测定的准确度更高。ZetaView的浓度测量值（% CV范围：0.0-4.7）也比NanoSight NS 300的测量值（% CV范围：5.4-10.7）更精确。相反，定量TEM成像表明，与ZetaView（% D-TEM范围：111.8-205.7）相比，NanoSight NS 300（% D-TEM范围：79.5-134.3）的EV尺寸更准确，同时具有同等的可重复性（NanoSight NS 300% CV范围：0.8-6.7; ZetaView：1.4-7.8）。然而，与TEM和SP-IRIS相比，两种装置均未能报告低于60 nm的峰值EV直径。总之，NTA设备在影响测量结果的硬件和软件方面存在很大差异。ZetaView提供了更准确和可重复的EV浓度描述，而NanoSight NS 300提供了更高分辨率的尺寸测量。
The expanding field of extracellular vesicle (EV) research needs reproducible and accurate methods to characterize single EVs. Nanoparticle Tracking Analysis (NTA) is commonly used to determine EV concentration and diameter. As the EV field is lacking methods to easily confirm and validate NTA data, questioning the reliability of measurements remains highly important. In this regard, a comparison addressing measurement quality between different NTA devices such as Malvern's NanoSight NS300 or Particle Metrix' ZetaView has not yet been conducted. To evaluate the accuracy and repeatability of size and concentration determinations of both devices, we employed comparative methods including transmission electron microscopy (TEM) and single particle interferometric reflectance imaging sensing (SP-IRIS) by ExoView. Multiple test measurements with nanospheres, liposomes and ultracentrifuged EVs from human serum and cell culture supernatant were performed. Additionally, serial dilutions and freeze-thaw cycle-dependent EV decrease were measured to determine the robustness of each system. Strikingly, NanoSight NS300 exhibited a 2.0-2.1-fold overestimation of polystyrene and silica nanosphere concentration. By measuring serial dilutions of EV samples, we demonstrated higher accuracy in concentration determination by ZetaView (% BIAS range: 2.7-8.5) in comparison with NanoSight NS300 (% BIAS range: 32.9-36.8). The concentration measurements by ZetaView were also more precise (% CV range: 0.0-4.7) than measurements by NanoSight NS300 (% CV range: 5.4-10.7). On the contrary, quantitative TEM imaging indicated more accurate EV sizing by NanoSight NS300 (% D-TEM range: 79.5-134.3) compared to ZetaView (% D-TEM range: 111.8-205.7), while being equally repeatable (NanoSight NS300% CV range: 0.8-6.7; ZetaView: 1.4-7.8). However, both devices failed to report a peak EV diameter below 60 nm compared to TEM and SP-IRIS. Taken together, NTA devices differ strongly in their hardware and software affecting measuring results. ZetaView provided a more accurate and repeatable depiction of EV concentration, whereas NanoSight NS300 supplied size measurements of higher resolution.