Formvar carbon coated 400 mesh copper grid

Phages were prepared for TEM using a modification of the method described by Fortier and Moineau (35 (link)). Prior to observation, 1.5 ml of crude lysate was centrifuged for 1 h at 4°C and 24,000 × g. A fraction of the supernatant (approximately 1.4 ml) was gently removed and discarded, and 1 ml of ammonium acetate (0.1 M, pH 7.5) was added to the remaining lysate, which was then centrifuged as described above. This step was performed twice. Washed phage samples were visualized by negative-stain transmission electron microscopy. A glow-discharged Formvar/carbon-coated 400-mesh copper grid (Ted Pella, Inc., Redding, CA) was floated on a 25-μl droplet of the sample suspension for 5 min and transferred quickly to 2 drops of deionized water, followed by transfer to a droplet of 2% aqueous uranyl acetate stain for 30 s. The grid was blotted with filter paper and air dried. Samples were observed using a JEOL JEM-1230 transmission electron microscope operating at 80 kV (JEOL USA, Peabody, MA), and images were taken using a Gatan Orius SC1000 charge-coupled-device camera with Gatan Microscopy (suite 3.0) software (Gatan, Inc., Pleasanton, CA).

Bodo saltans lysates after BsV infection were applied to the carbon side of a formvar-carbon coated 400 mesh copper grid (TedPella, CA) and incubated at 4°C in the dark overnight in the presence of high humidity. Next, the lysate was removed and the grids were stained with 1% Uranyl acetate for 30 s before observation on a Hitachi H7600 transmission electron microscope at 80 kV.

Lysates of Spumella elongata after infection with Chromulinavorax destructans were applied to the carbon side of a formvar carbon-coated 400-mesh copper grids (TedPella, CA, USA) and incubated at 4°C in the dark overnight in the presence of high humidity. Next, the lysate was removed and the grids were stained with 1% Uranyl acetate for 30 s.