Biomedical Physics Division, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, 5 Pasteura Street, 02-093 Warsaw, Poland.
1st Department of Clinical Radiology, Medical University of Warsaw, 5 Chałubińskiego Street, 02-004 Warsaw, Poland.
Int J Mol Sci. 2022 Sep 3;23(17):10074. doi: 10.3390/ijms231710074.
Extracellular vesicles are evaluated by nanoparticle tracking analysis (NTA), providing information on their hydrodynamic diameters, and by atomic force microscopy (AFM) to calculate their geometric diameters. The aim of this study is to explore the influence of Brownian movements in a sample drop and preparation time on imaging-based measurements and to determine the relationship between the geometric and hydrodynamic sizes of the extracellular vesicles measured by the AFM and the NTA, respectively. Exosomes derived from the human prostate cancer cell line PC3 were evaluated by NTA and AFM, and those results were compared with Monte Carlo simulations. The mean size, evaluated by AFM shortly after application on the mica substrate, is less than its real value. It obtains the correct value faster for a thinner sample drop. Fitting the log-normal distribution to the geometric and hydrodynamic diameters leads to the conclusion that the latter could arise from the former by linear scaling by a factor that could be used to characterize the analyzed extracellular vesicles. The size of the vesicles attached to the mica substrate depends on time. The effect of Brownian motion and stretch of the lipid bilayer should be considered in the context of exosome AFM studies.
细胞外囊泡可通过纳米颗粒跟踪分析（NTA）评估，提供有关其水动力直径的信息，并通过原子力显微镜（AFM）计算其几何直径。本研究旨在探讨样品滴和制备时间的布朗运动对基于成像的测量的影响，并确定 AFM 和 NTA 分别测量的细胞外囊泡的几何尺寸和水动力尺寸之间的关系。用人前列腺癌细胞系 PC3 衍生的外泌体进行 NTA 和 AFM 评估，并将这些结果与蒙特卡罗模拟进行比较。在云母衬底上应用后不久，通过 AFM 评估的平均尺寸小于其真实值。对于更薄的样品滴，它可以更快地获得正确的值。将对数正态分布拟合到几何和水动力直径，得出后者可以通过线性比例因子从前者得出的结论，该因子可用于表征分析的细胞外囊泡。附着在云母衬底上的囊泡的大小取决于时间。在进行外泌体 AFM 研究时，应考虑布朗运动和脂质双层的拉伸对囊泡尺寸的影响。
