Tecnai g20 electron microscope

The surface morphologies of the samples were characterized by scanning electron microscopy (SEM; Quanta 200; FEI, USA). The surface chemical compositions of the samples were determined via energy dispersive spectroscopy (EDX, attached to the SEM). Transmission electron microscopy (TEM) experiments were performed on a Tecnai G₂₀ electron microscope (FEI, USA) with an acceleration voltage of 200 kV. Carbon-coated copper grids were used as the sample holders. The crystalline structures of the samples were identified by X-ray diffraction (XRD; D/MAX 2200; Rigaku, Japan) using Cu K_α radiation (λ = 1.5418 Å) at a 2θ scan rate of 4° min⁻¹, 40 kV, 40 mA, ranging from 5° to 80°. The presence of functional groups in the samples was confirmed through Fourier-transform infrared (FTIR) spectroscopy (Spectrum One; PerkinElmer, USA). The surface elemental composition analyses were conducted based on X-ray photoelectron spectroscopy (XPS; Scientific-K-Alpha 1063; Thermo Fisher, UK) with an Al Kα monochromatic X-ray source, in which all of the binding energies were calibrated with reference to the C 1s peak (284.8 eV).

Cells were grown to 90% confluence before being washed, scraped, pelleted, and fixed in 2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer (pH 7.2) overnight. Samples were post fixed in 1% osmium tetroxide for 2 h, stained in bloc with 0.5% uranyl acetate, rinsed, and dehydrated in graded ethanol. After immersion in propylene oxide, samples were embedded in epoxy resin (Spurr, Electron Microscopy Sciences, EMS, Hatfield PA, USA) and polymerized for 42 h at 75 °C. Ultrathin sections were stained with lead citrate and uranyl acetate and examined in a FEI Tecnai G20 Electron microscope at 200 kVA.

The exfoliation mixture was first centrifuged at 2500 rpm to remove large graphite particles and possible planktonic microorganisms, and then the supernatant was filtered through a 0.1 μm Durapore PVDF filter (Millipore Co.,). The remaining solid materials on the filter were washed with the following sequence: 18 MΩ water, 80% ethanol, 18 MΩ water, 1 N HCl, and 18 MΩ water again. The resulting materials called BEGO were re-suspended in 18 MΩ water for characterization. UV-vis absorption spectra were recorded using a Thermo GENESYS 10S UV-vis spectrophotometer. Fourier transform infrared spectroscopy (FT-IR) was performed by using a Thermo-Nicolet FT-IR Avatar 370 spectrometer. XPS analysis was carried out by a Kratos Axis His spectrometer. Raman spectra were obtained by Raman Microscope with wavelength of 532 nm and a 100 × objective. Atomic force microscopy (AFM) images were generated on a VEECO Dimension 3100. Transition electron microscopy (TEM) and electron diffraction images were obtained using a FEI Tecnai G20 electron microscope. Samples for AFM and TEM imaging were prepared by drop-casting the dispersion onto freshly cleaved mica substrates and lacey carbon TEM grid, respectively, which were then air dried under ambient lab conditions.

We used protocols similar to previous studies (Andreani et al., 2016 (link), 2017 (link)) for embedding and transmission electron microscopy. The only difference was the infection time points at which the samples were collected. The time points were 0, 4, 8, 12, 16, 20, 24, and 36h post-infection. Microscopy was performed using a Tecnai G20 electron microscope (FEI, Germany) operating at 200keV, and the size of the particles was measured using ImageJ.¹