How to apply FLUCS in single cells and living embryos

This protocol is associated to the publication "Non-invasive perturbations of intracellular ow reveal physical principles of cell organization" by Mittasch et al., published in Nature Cell Biology.In the following, we describe the procedure for maintenance and preparation of C. elegans zygotes for imaging and application of FLUCS.Preparation of S. cerevisiae cells for microscopy and intracellular ow perturbations.S. cerevisiae cells were grown into logarithmic stage.Subsequently the S. cerevisiae cells were either grown for 2 hours or alternatively energy depleted through addition of 2-Desoxy-D-Glucose \ (inhibition of glycolysis) and antimycin A \(inhibition of mitochondrial ATP production) for 2 h prior to ow induction.This treatment causes a more than 95% reduction in cellular ATP 2. Potentially this protocol is suitable to energy deplete all S. cerevisiae strains and closely related yeast species.A speci c marker such as µNS-GFP1is needed to observe subsequent ow perturbations by microscopy.