Visualizing Bacterial Infection via SEM and TEM

SEM and TEM were performed to confirm localization and view cell–cell interactions^{70 (link)}. For TEM, VK2 E6/E7 cells were grown in six-well tissue culture plates and infected with strain UTI89 at the indicated MOI as described for the adherence and invasion assay. At the indicated time point cells were washed, and fixed in 2.5% glutaraldehyde in 100 mM sodium cacodylate for 30 min at room temperature. Specimens were placed on glow-discharged formvar/carbon-coated copper grids and stained with 1% uranyl acetate for 90 s. A Philips/FEI T-12 transmission electron microscope was used for sample analysis. For SEM, VK2 E6/E7 cells were grown on glass cover slips in 12-well plates, infected with the bacteria for 2 h and fixed with 2.5% glutaraldehyde in 100 mM sodium cacodylate for 30 min at room temperature. Samples were critical point dried and placed onto aluminum stubs prior to sputter coating with gold-palladium. Images were collected for each sample and representative images were taken with a FEI Quanta 250 field-emission gun scanning electron microscope (Field Electron and Ion Company, Hilllsboro, OR). Images were taken from a minimum of three biological replicates.