Tecnai t12 electron microscope

High resolution TEM imaging (FEI, Hillsboro, USA) was performed using 120 kV Tecnai T12 electron microscope equipped with side-mounted Olympus VELITA (2 K × 2 K) CCD camera. GO-H₂O and GO-Met-H₂O coated carbon grids were imaged at 120 kV at different magnification range 4200×–63,000×. Similarly GO solution was directly deposited on Cu-grids without any carbon support films for negative staining purposes. Diffraction pattern of GO monolayer and multilayers were recorded at diffraction mode using T12 microscope. Furthermore, the GO-coated Cu-grids were analyzed with JEOL IT-300 (JEOL. Ltd., Tokyo, Japan) scanning electron microscopy. Data were collected at 30 kV high vacuum mode at 10,000× magnification using secondary electron detector (SED).

300 mesh Cu-grids without any carbon support film was selected to prepare TEM grids with GO-support film. GO-H₂O and GO-Met-H₂O solution was directly deposited on TEM grids without any carbon support film, and grids were dried at room temperature overnight. These GO coated grids were directly used for negative staining. Three different biological samples, E. coli 70S Ribosome (NEB P0763S), Vibrio cholerae Cytolysin and MdtB from E. coli were visualized using this GO-coated grids. Standard protocols were followed to prepare negative staining grids, except the glow discharge step before staining. Briefly, 3 µL samples were incubated for 30–60 s on the copper grids coated with methanol-water treated graphene oxide. Excess buffer and bio sample was blotted, and negative staining was executed using 2% uranyl acetate. The entire grids with biological samples were screened at 4200× low magnification to visualize the GO monolayer formation and stability in presence of biological sample and staining solution. Additionally, maximum grids squares of GO-coated grids were inspected at higher magnification (20,000×) to analyze the biological sample distribution on GO support film. The imaging was performed at room temperature using a 120 kV Tecnai T12 electron microscope at 120 kV at pixel size 2.54 Å using side-mounted Olympus VELITA (2K × 2K) CCD camera.