Label-free imaging of lipid-droplet intracellular motion in early Drosophila embryos using femtosecond-stimulated Raman loss microscopy.

Lipid droplets are complex organelles that exhibit highly dynamic behavior in early Drosophila embryo development. Imaging lipid droplet motion provides a robust platform for the investigation of shuttling by kinesin and dynein motors, but methods for imaging are either destructive or deficient in resolution and penetration to study large populations of droplets in an individual embryo. Here we report real-time imaging and quantification of droplet motion in live embryos using a recently developed technique termed "femtosecond-stimulated Raman loss" microscopy. We captured long-duration time-lapse images of the developing embryo, tracked single droplet motion within large populations of droplets, and measured the velocity and turning frequency of each particle at different apical-to-basal depths and stages of development. To determine whether the quantities for speed and turning rate measured for individual droplets are sufficient to predict the population distributions of droplet density, we simulated droplet motion using a velocity-jump model. This model yielded droplet density distributions that agreed well with experimental observations without any model optimization or unknown parameter estimation, demonstrating the sufficiency of a velocity-jump process for droplet trafficking dynamics in blastoderm embryos.