单个腺病毒颗粒的原子力显微镜液相成像及样品制备方法

Sample Preparation and Imaging of Single Adenovirus Particle Using Atomic Force Microscopy in Liquid

原子力显微镜(Atomic force microscopy,AFM)作为一种纳米级高分辨率的探针扫描显微镜,在病毒形态学研究等领域有广泛的应用。为更好地利用AFM在接近病毒生理环境(液相)中观测单个病毒颗粒,选择合适的基底吸附病毒样品是重要的前提保障。本文介绍了一种病毒样品制备及AFM液相成像方法。首先制备疏水化处理的玻璃盖玻片作为吸附基底,再将观察样品腺病毒重组载体颗粒(Adenovirus type 5F35,Ad5F35)吸附于基底上,建立在液相下单个病毒颗粒的原子力显微镜成像方法。通过该方法获取了单个腺病毒颗粒在液相下的高分辨率纳米级高度图(height image),且病毒颗粒保持了约90nm的正确测量高度。实验结果表明疏水化处理的玻璃基底具有良好的吸附力,有助于病毒颗粒牢固地吸附在基底上,且不会引起较大的形变。本研究为AFM观测病毒等生物样品提供了一种较为合适的吸附基底和在液相中的AFM成像方法,对后期开展病毒形态结构、物理属性及其与宿主细胞受体相互作用的研究打下基础。

Atomic force microscopy（AFM）,as a sophisticated imaging tool with nanoscale resolution,is widely used in virus research and the application of functional viral particles.To investigate single viruses by AFM in a physiologically relevant environment（liquid）,an appropriate surface treatment to properly adhere the viruses to the substrate is essential.Here we discuss hydrophobic treated glass coverslips as a suitable substrate for the adhesion of single adenovirus particle（Adenovirus type 5 F35,Ad5F35） when studied with AFM in liquid.From the high resolution AFM images,the orientation of the adhered virus particles can be distinguished.Furthermore,the particles exhibit the expected height of ~90 nm.This illustrates that the viruses adhere to the substrate firmly without large deformations.Hence,the described method works well on（fragile） viruses.The described experimental approach can be widely used for AFM studies in liquid of virus structure and mechanics as well as for investigating the interaction of viruses with cellular receptors.