A New Label-Free Technique for Analysing Evaporation Induced Self-Assembly of Viral Nanoparticles Based on Enhanced Dark-Field Optical Imaging

Nanoparticle self-assembly is a complex phenomenon, the control of which is complicated by the lack of appropriate tools and techniques for monitoring the phenomenon with adequate resolution in real-time. In this work, a label-free technique based on dark-field microscopy was developed to investigate the self-assembly of nanoparticles. A bio-nanoparticle with complex shape (T4 bacteriophage) that self-assembles on glass substrates upon drying was developed. The fluid flow regime during the drying process, as well as the final self-assembled structures, were studied using dark-field microscopy, while phage diffusion was analysed by tracking of the phage nanoparticles in the bulk solutions. The concentrations of T4 phage nanoparticles and salt ions were identified as the main parameters influencing the fluid flow, particle motion and, consequently, the resulting self-assembled structure. This work demonstrates the utility of enhanced dark-field microscopy as a label-free technique for the observation of drying-induced self-assembly of bacteriophage T4. This technique provides the ability to track the nano-sized particles in different matrices and serves as a strong tool for monitoring self-assembled structures and bottom-up assembly of nano-sized building blocks in real-time.