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3.5 Morphological test of the bacterial cells Scanning electron microscopy (SEM) was used to examine morphological changes of the bacterial cells before and after treating with ZnO NPs and Ag NPs. The three bacterial strains treated or untreated with NPs were fixed with a primary fixative (2.5 glutaraldehyde, 2 % paraformaldehyde in 0.1 M Na-Cacodylate buffer, pH 7.4). The samples were then rinsed three times with ultrapure water, followed by dehydration with a series of ethanol solutions (10, 30, 50, 70, 90, and 100%). The dehydrated samples were immediately dried by a critical point dryer (Auto-Samdri 815 Automatic Critical Point Dryer; Tousimis, Rockville, MD, USA), mounted on SEM stubs and coated with a thin layer of carbon using a sputter coater (K575X Turbo Sputter Coater; Emitech, Ltd, Kent, UK). The coated samples were observed under SEM (FEI Quanta 600, FEI Company, Hillsboro, OR, USA). Transmission electron microscopy (TEM) was used to characterize the size of the NPs and to observe the morphology of bacterial cells after treatment with ZnO NPs and Ag NPs. Three samples of bacterial cells treated with and without NPs were fixed with a primary fixative and microwaved under vacuum conditions in a Pelco Biowave (Ted Pella, Inc., Redding, CA, USA) at 120 W. The samples were rinsed with 0.1 M cacodylate buffer and embedded in histogel, followed by a secondary microwave fixation with a buffered (0.1 M cacodylate, 0.01 M of 2-mercaptoethanol, and 0.13 M of sucrose) 1% osmium tetroxide. The samples were then quickly rinsed three times with 0.1M 2- mercaptoethanol, 0.13M of sucrose and then rinsed three times with ultrapure water. Then, samples were dehydrated with ethanol solutions (20, 50, 70, 90, and 100%) and 21PDF Image | ZINC OXIDE AND SILVER NANOPARTICLES ON INTESTINAL BACTERIA
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