Incubation samples of the FUS RRM domain at 25 °C at a protein concentration of 40 µM were imaged at 3, 6 and 10 days of the incubation in 1 mM phosphate buffer (pH 6.8), by a TEM microscope (Jeol Jem 2010f Hrtem, Japan) operating at an accelerating voltage of 200 kV as previously described24 (link).
Briefly, for EM imaging, a 5 µl aliquot of the incubation or aggregate solutions was placed onto the Cu grids (coated with carbon film; 150 mesh; 3 mm in diameter) and negatively stained with 5 μl of 2% neutral, phosphotungstic acid (PTA). This aliquot was allowed to settle on Cu grid for 30 sec before the excess fluid was drained away. The Cu grid was later air-dried for another 15 mins before being imaged.
Briefly, for EM imaging, a 5 µl aliquot of the incubation or aggregate solutions was placed onto the Cu grids (coated with carbon film; 150 mesh; 3 mm in diameter) and negatively stained with 5 μl of 2% neutral, phosphotungstic acid (PTA). This aliquot was allowed to settle on Cu grid for 30 sec before the excess fluid was drained away. The Cu grid was later air-dried for another 15 mins before being imaged.
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