Samples were prepared for EM as previously described (46 (link)). In brief, 5-µl aliquots of samples were absorbed to a glow-discharged 400-mesh copper grid covered with carbon-coated collodion film. Grids were washed in four drops of water and then stained with two drops of uranyl formate (0.75%). Samples were imaged on a Morgagni electron microscope (FEI, Hillsboro, OR) operated at an acceleration voltage of 100 kV and equipped with a 1kx1k charge-coupled-device (CCD) camera (ATM). Images were recorded at a magnification of ×28,000. Images used to generate class averages were collected using a TF20 electron microscope (FEI, Hillsboro, OR) equipped with a field emission gun with an acceleration voltage of 200 kV under low-dose conditions at ×62,000 magnification and a defocus value of ~1.5 µm on a 4kx4k Gatan Ultrascan CCD camera.
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Transmission Electron Microscopy Sample Preparation
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Other organizations : Vanderbilt University, VA Tennessee Valley Healthcare System
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independent variables
- Preparation method for EM samples: 5-µl aliquots of samples were absorbed to a glow-discharged 400-mesh copper grid covered with carbon-coated collodion film, washed in four drops of water, and then stained with two drops of uranyl formate (0.75%)
- Morphological characteristics of samples observed under electron microscope
- Acceleration voltage of electron microscope (100 kV for Morgagni, 200 kV for TF20)
- Magnification of electron microscope images (×28,000 for Morgagni, ×62,000 for TF20)
- Defocus value (~1.5 µm) for TF20 electron microscope
- No positive or negative controls were explicitly mentioned in the provided information.
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