Carbon formvar copper grid

It was performed on the microscopy platform IBiSA at the Institut Curie, Orsay, France. U87 cells were plated on microscopic glass slides. The samples were incubated with 1 mM GdBN during 1 hour. The slides were then rinsed with PBS 1X and fixed with a mixture of 2.5% glutaraldehyde and 4% paraformaldehyde diluted in PBS 1X. After rinsing with PBS 1X, the cells were dehydrated using ethanol in gradient concentrations and embedded step by step in Epon resin. After resin polymerisation, the samples were cut using an ultramicrotome in 100 nm-thick slices. The ultrathin sections were deposited on carbon-formvar copper grids (Agar scientific) and observed under Z-loos mode (10 eV window) in a JEOL 2200FS electron microscope operated at 200 kV. Measurements were performed for close to 20 U87 cells of 4 different slices. Additional Electron Energy Loss Spectroscopy (EELS) measurements were performed using an omega-filter.

To analyse percentages of piliated cells, each HI strain was back-diluted and grown to OD₆₀₀ 0.1–0.5 in PY broth at 29°C 200 rpm. Cells were then stained with 2.0% phosphotungstic acid (PTA) on carbon formvar copper grids (Agar Scientific) and analysed for the presence/absence of a pilus structure, as described previously [26] (link).

Electron microscopy was performed to examine the presence of flagella on the cell surface of the C. difficile strains. C. difficile cells from mid-exponential cultures were diluted in sterile water and then adsorbed onto carbon formvar copper grids (Agar Scientific Ltd., UK) for 5 min. After air-drying overnight, the grid was quickly washed once with sterile distilled water for 10 s. The air-dried grid was visualized using a JOEL JEM1010 transmission electron microscope operating at 80 kV.