Microfluidic Device for High-Resolution Cytoskeleton Imaging and Washout Assays in Physcomitrium (Physcomitrella) patens

Visualizing cytoskeleton dynamics at high spatiotemporal resolution provides valuable insights into the way the dynamics change as well as its interactions with multiple proteins in order to maintain cellular function. Oblique illumination fluorescent microscopy is a popular technique to image cellular events localized near the plasma membrane. In this chapter, we provide detailed protocols for high-resolution cytoskeleton imaging using protonema and gametophore cells of the moss Physcomitrella (Physcomitrium patens) in the microfluidic device. These include preparation of the polydimethylsiloxane (PDMS) device, culture of moss cells, and both short- and long-term oblique illumination fluorescent microscopy. We also describe how to introduce to, and wash out from, the device chemical compounds, such as microtubule-disrupting drugs, during live-cell imaging.