Spatial light modulation for interferometric scattering microscopy.

Interferometric scattering (iSCAT) microscopy enables high-speed and label-free detection of individual molecules and small nanoparticles. Here we apply point spread function engineering to provide adaptive control of iSCAT images using spatial light modulation. With this approach, we demonstrate improved dynamic spatial filtering, real-time background subtraction, focus control, and signal modulation based on sample orientation.

Real-Time Monitoring and Control of Nanoparticle Formation.

Methods capable of controlling synthesis at the level of an individual nanoparticle are a key step toward improved reproducibility and scalability in engineering complex nanomaterials. To address this, we combine the spatially patterned activation of the photoreductant sodium pyruvate with interferometric scattering microscopy to achieve fast, label-free monitoring and control of hundreds of gold nanoparticles in real time. Individual particle growth kinetics are well-described by a two-step nucleation-autocatalysis model but with a distribution of individual rate constants that change with reaction conditions.

Measuring Encapsulation Efficiency in Cell-Mimicking Giant Unilamellar Vesicles.

One of the main drivers within the field of bottom-up synthetic biology is to develop artificial chemical machines, perhaps even living systems, that have programmable functionality. Numerous toolkits exist to generate giant unilamellar vesicle-based artificial cells. However, methods able to quantitatively measure their molecular constituents upon formation is an underdeveloped area.

Single-molecule imaging of pore-forming toxin dynamics in droplet interface bilayers.

Single-channel recording from pore-forming toxins (PFTs) provides a clear and direct molecular readout of toxin action. However to complete any mechanistic understanding of PFT behavior, this functional kinetic readout must be linked to the underlying changes in toxin structure, binding, conformation, or stoichiometry. Here we review how single-molecule imaging methods might be used to further our understanding of PFTs, and provide detailed practical guidance on the use of droplet interface bilayers as a method capable of examining both single-molecule fluorescence and single-channel electrical signals from PFTs.