Diaphot inverted phase contrast microscope

Device dimensions were chosen to ensure that cells would be retained in the culture well despite multiple fluid replacements as previously described^{17 (link)}. Briefly, by having a wide culture well and allowing the cells to settle to the bottom of the well out of the direct path of fluid flow, shear stresses on the cells were minimized. To operate the devices, 5 μL spore suspension at 1 × 10⁷ spores/mL or 5 μL yeast suspension at 1 × 10⁶ yeast/mL was loaded into the input port and allowed to passively pump through the device. Cells were allowed to settle to the bottom of the well for 5 minutes before 5 μL 2X germination medium was added for a final concentration in the device of 1X (as demonstrated in Supplemental Figure 1, a volume replacement displaces half of the liquid in a device). Unless stated otherwise, germination or yeast growth was assessed after 16 hours of incubation at 30°C. All assays were performed in triplicate. At assay points, two images were taken per device. Light microscopy was carried out using a Nikon Diaphot inverted phase contrast microscope fitted with a 20X objective and Nikon D5000 camera.