High-Throughput Pulsed Stimulated Brillouin Microscopy for Cell Biomechanics - ABSTRACT The mechanical properties of cells are critically involved in the regulation of numerous biological processes. This has spurred the flourishing of mechanical testing methods for biomedical and biomaterial research in the past three decades. Current instruments, though, require contact, are invasive, or are limited to the global analysis of a sample. The goal of this proposal is the development of a novel optical microscopy modality for cell biomechanics, based on pulsed Stimulated Brillouin scattering (SBS), which will be non-contact, to probe different experimental settings beyond 2D cultures; non-perturbative, since cells are known to react to mechanical stimuli; and, high-resolution, since biological cells are not homogeneous. In the past few years, Brillouin microscopy has rapidly emerged, and its potential has been established in ophthalmology, tumor biology and development biology. However, Brillouin microscopy has yet to reach widespread adoption for two main challenges: acquisition speed still limited to ms/pixel and mechanical interpretation of the extracted signatures. Based on strong preliminary data, here we plan to overcome these challenges with the development of novel laser source and detection scheme for rapid SBS and the development of co-located maps of refractive index and density to extract accurate mechanical information. In Aim 1, we will implement our technical innovations to maximize the throughput of SBS microscopy, with an expected 100-fold improvement in acquisition speed compared with current state-of-the-art. In Aim 2 we will validate the modulus estimation from SBS characterization against gold-standard methods after uniquely factoring direct measurements of refractive index and density. In Aim 3, we will field-test the high-speed SBS microscope for cell migration experiments where biomechanics is known to be crucial but other mechanical testing methods are not feasible. This will both provide quantitative feedback of performances to enable fine-tuning of the instrument development and demonstrate the technology in settings that are familiar to the biological research community to facilitate communication of our instrument features and adoption of our technology by the intended final users.
