S 2600 vp sem

The morphology of P. yuhuli AB48 was described using light and scanning electron microscopy (SEM) techniques. For brightfield images, a Zeiss Axio Observer 7 microscope with an attached Calibiri 7 light source and an Axiocam 702 mono camera was used. For the detection of autofluorescence, a mRF12 filter was used. The samples imaged using SEM were isolated from suspension and filtered onto 0.2 µm Nucleopore filters where they were fixed with 4% formaldehyde/2% glutaraldeyde in 0.05 M, pH 7.4 sodium cacodylate. The cells were washed 3x with fixative-free buffer, then post-fixed with 1% buffered OsO₄ before multiple washes with distilled, deionized water. Following the washes, the samples were stacked in a stainless-steel filter holder, and taken through a staged, microwave dehydration using 1 min at power-level 3 (∼180W) in 30, 50, and 70% ethanol. The samples were kept at 70% ethanol overnight at −20°C. Following overnight dehydration, the samples were treated for 1 min PL3 at 80, 90, and 95% ethanol and finally for 3 min PL3 at 100% dry ethanol for dehydration, twice. The final dehydration step was performed for 10 min at room temperature in 100% dry ethanol. The samples were then critically point-dried (Tousimis Auto-Sam Dri 815B) mounted to aluminum SEM stubs using double-sided tabs, sputter coated with 10 nm gold (Cressington 208HR), and imaged on a Hitachi S2600 VP-SEM.