Long-Term High-Resolution Imaging of Developing C. elegans Larvae with Microfluidics

Abstract

Long-term studies of Caenorhabditis elegans larval development traditionally require tedious manual observations because larvae must move to develop,and existing immobilization techniques either perturb development or are unsuited for young larvae. Here, we present a simple microfluidic deviceto simultaneously follow development of ten C.elegans larvae at high spatiotemporal resolution from hatching to adulthood (∼3days). Animals grownin microchambers are periodically immobilized bycompression to allow high-quality imaging of even weak fluorescence signals. Using the device,we obtain cell-cycle statistics for C.elegans vulval development, a paradigm for organogenesis. Wecombine Nomarski and multichannel fluorescence microscopy to study processes such as cell-fate specification, cell death, and transdifferentiation throughout post-embryonic development. Finally, we generate time-lapse movies of complex neural arborization through automated image registration. Our technique opens the door to quantitativeanalysis of time-dependent phenomena governing cellular behavior during C.elegans larval development.