Nikitin Nikolai, Trifonova Ekaterina, Evtushenko Evgeniy, Kirpichnikov Mikhail, Atabekov Joseph, Karpova Olga
Lomonosov Moscow State University, Moscow, Russia.
PLoS One. 2015 Nov 6;10(11):e0142415. doi: 10.1371/journal.pone.0142415. eCollection 2015.
Now, as before, transmission electron microscopy (TEM) is a widely used technique for the determination of virions size. In some studies, dynamic light scattering (DLS) has also been applied for this purpose. Data obtained by different authors and using different methods could vary significantly. The process of TEM sample preparation involves drying on the substrate, which can cause virions to undergo morphology changes. Therefore, other techniques should be used for measurements of virions size in liquid, (i.e. under conditions closer to native). DLS and nanoparticle tracking analysis (NTA) provide supplementary data about the virions hydrodynamic diameter and aggregation state in liquid. In contrast to DLS, NTA data have a higher resolution and also are less sensitive to minor admixtures. In the present work, the size of non-enveloped icosahedral viruses of different nature was analyzed by TEM, DLS and NTA: the viruses used were the encephalomyocarditis virus (animal virus), and cauliflower mosaic virus, brome mosaic virus and bean mild mosaic virus (plant viruses). The same, freshly purified, samples of each virus were used for analysis using the different techniques. The results were compared with earlier published data and description databases. DLS data about the hydrodynamic diameter of bean mild mosaic virus, and NTA data for all examined viruses, were obtained for the first time. For all virus samples, the values of size obtained by TEM were less than virions sizes determined by DLS and NTA. The contribution of the electrical double layer (EDL) in virions hydrodynamic diameter was evaluated. DLS and NTA data adjusted for EDL thickness were in better agreement with TEM results.
如今，和以往一样，透射电子显微镜（TEM）是一种广泛用于确定病毒粒子大小的技术。在一些研究中，动态光散射（DLS）也被用于此目的。不同作者使用不同方法获得的数据可能会有显著差异。TEM样品制备过程包括在基质上干燥，这可能会导致病毒粒子发生形态变化。因此，应该使用其他技术来测量液体中病毒粒子的大小（即在更接近天然的条件下）。DLS和纳米颗粒跟踪分析（NTA）提供了关于病毒粒子在液体中的流体动力学直径和聚集状态的补充数据。与DLS相比，NTA数据具有更高的分辨率，并且对微量杂质不太敏感。在本研究中，通过TEM、DLS和NTA分析了不同性质的无包膜二十面体病毒的大小：所使用的病毒是脑心肌炎病毒（动物病毒）、花椰菜花叶病毒、雀麦花叶病毒和菜豆轻花叶病毒（植物病毒）。每种病毒相同的新鲜纯化样品用于使用不同技术进行分析。将结果与早期发表的数据和描述数据库进行了比较。首次获得了菜豆轻花叶病毒流体动力学直径的DLS数据以及所有检测病毒的NTA数据。对于所有病毒样品，通过TEM获得的大小值小于通过DLS和NTA确定的病毒粒子大小。评估了双电层（EDL）在病毒粒子流体动力学直径中的贡献。针对EDL厚度进行调整后的DLS和NTA数据与TEM结果更吻合。
