Formvar carbon coated copper grid 100 mesh

The morphology of the nanoemulsion droplets was studied using TEM. Briefly, 5 µL of the sample was placed on a formvar-carbon-coated copper grid (100 mesh, Electron microscopy sciences, Hatfield, PA, USA). Thereafter, the grid was rinsed twice with milli Q water and stained with 2% uranyl acetate solution (Ladd Research Industries, Williston, VT, USA), and any excess solution was removed with Whatman 3MM filter paper. The grid was then left to air-dry for a few minutes and imaged at 20,000× magnification using FEI Tecnai Spirit TWIN TEM (FEI, Hillsboro, OR, USA) operated at 120 kV voltage.

Food-grade TiO₂ (E171, Pretiox AV01PhG) was kindly provided by Giusto Faravelli S.p.A. (Milan, Italy). The product was 99.3% pure TiO₂ anatase, complying with the standard regulations of the European and US Pharmacopoeia, the Food and Drug Administration (FDA), and the Food Additive Regulations. To investigate whether serum could affect the morphology and colloidal properties of E171 over time, TEM analysis was performed, incubating 1 mg/100 µL of E171 in either H₂O or murine serum for 1, 4, or 24 h. To avoid any effects on the particle characteristics that would not occur in realistic conditions of use, the samples were not sonicated. From each sample, 5 μL were extracted, placed into a formvar/carbon-coated copper grid (100 mesh) (Electron Microscopy Sciences, Washington, PA, USA), and air-dried at room temperature. NP images were obtained with an energy-filtered transmission electron microscope (EFTEM, ZEISS LIBRA^® 120, Milano, Italy) coupled with an yttrium aluminum garnet (YAG) scintillator slow-scan CCD camera (Sharp eye, TRS, Moorenweis, Germany) with a focus aid option, enabling us to reach the precise eucentric plane in the observation of each single NP [21 (link)].