Us4000 ccd camera

Bacterial cells or purified NCs were applied to glow-discharged carbon-coated Cu grids and stained with 2% uranyl acetate. Images were acquired with Digital Micrograph (Gatan Inc.) on a FEI Tecnai T12 electron microscope (120 kV) equipped with a US4000 CCD camera (Gatan Inc.).

3 μl of sample was applied to a freshly glow-discharged carbon-coated grid. Sample was incubated for 1 min at room temperature; excess sample was removed by blotting off with the filter paper and stained with 0.75% (w/v) of uranyl formate (UF). Grids were imaged using Tecnai T12 transmission electron microscope operated at a voltage of 120 kV. Images were recorded at a magnification of 50,000X and a defocus value of-1.0 μm on a Gatan US4000 CCD camera. All images were binned over 2 × 2 pixels to obtain a pixel size of 4.16 Å on specimen level.
For DNA origami, 1,421 particles were picked manually from the binned micrographs by using boxer of the EMAN 1.9 software suite [11 (link)]. Particles were then subjected to reference-free 2D classification using EMAN 1.9 image processing suite [11 (link)] and classified into 10 classes.

250 nm thick sections were collected on Formvar-coated slot grids for TEM tomography. Next, 15 nm colloidal gold particles (Sigma-Aldrich, Saint-Louis, MO, USA) were applied on the supporting film to serve as fiducial markers for tomogram alignment. The tomographic data collection was performed using Tecnai F30 transmission electron microscope (Thermo Fisher Scientific, Waltham, MA, USA) equipped with a US4000 CCD camera (Gatan, Pleasanton, CA, USA). Regions of interests (ROIs) were pre-exposed at 3000 e/Ų to minimize the section drift and warping [56 (link),57 (link)]. The dual-axis tomographic acquisition was done at 12,000× magnification (1.96 nm/pixel, camera binning 2) using Serial EM software (University of Colorado [55 (link)]) running a continuous tilt-scheme from −60° to +60° and a constant angular increment of 1.5°. Tilt series alignment and tomogram reconstruction were performed using ETOMO program of IMOD software package [58 (link)].

To determine the shape of exosomes in maternal plasma, 3 μL of prepared exosome suspension was pipetted onto a copper grid with quantifoil support film (QUANTIFOIL, Germany). The support film was patterned with a regular array of circular holes. When the excess liquid was blotted away from the grid, a 60–120 nm-thick film of sample suspension remained in these holes. The grid was then plunged into a small crucible of liquid ethane that was cooled to its melting point by liquid nitrogen. In the liquid ethane, the sample suspension was cooled at over 10,000 degrees/s, solidifying the water in an amorphous state. This “vitrification” process preserves the exosomes in their native state without distorting their geometry by crystallization and density change^{41 (link)}. The vitrified sample on the grid was then placed in a Gatan 626 specimen holder (Gatan, Pleasanton, CA), which was placed in a JEOL 2100 TEM (JEOL, Osaka, Japan). The samples were imaged with a 200 kV electron beam from a LaB6 emission source, and images were recorded on a Gatan US4000 CCD camera. Images were captured at 15,000–30,000 magnification.