Abstract
Self-assembly is widely used by biological systems to build complex and functional nanoscale structures, but the assembly pathways are difficult to observe because of the small length scales and wide range of time scales involved. We describe a label-free optical method to directly probe the assembly of individual nanostructures, and we apply it to measure the kinetics of assembly of viral capsids 28 nm in diameter and consisting of 90 protein subunits. We use a rigid flow cell to inject a solution of the coat protein of bacteriophage MS2 over multiple MS2 RNA strands that are tethered to a microscope coverslip by specific DNA linkages. Using an interferometric detection scheme, we measure changes in the intensity of light scattered from the proteins while they are assembling around each RNA. The low photodamage afforded by elastic scattering enables high illumination intensities and temporal resolutions down to 1 ms, while 3D-active stabilization of the microscope extends the measurement duration to 600 s or longer. With this wide range of timescales, we find that the assembly is characterized by an exponential distribution of wait times preceding a rapid growth phase, suggesting that the pathway under the conditions we investigate is nucleation followed by growth. Because the method can measure the assembly of many individual capsids in parallel, from start to finish, it offers a direct view of the self-assembly process not accessible to bulk scattering or spectroscopic techniques. It can be adapted to study the assembly of other viruses, biomolecular assemblies, and synthetic nanostructures.
Footnotes
↵E-mail: vnm{at}seas.harvard.edu