Aggregated proteins from the microfluidic device were obtained by excising the PDMS block from the glass slides using a sterile scalpel and flushing the surfaces repeatedly with 100 μl of ultrafiltered deionized water. The collected liquid was refrigerated along with the remaining liquid in the reservoirs until further analysis. Procedures for collecting aggregated proteins from shaking experiments as well as for EM sample preparation were described previously7 (link). Negative stain EM data were collected on a JEOL JEM1400 electron microscope with a Gatan Orius 832 CCD. Cryo-EM data were collected on a JEOL JEM-2100F microscope with K2 Summit direct detector. Data were processed using EMAN259 (link) and RELION-445 (link) software packages.
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Microfluidic Protein Aggregation Analysis
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Microfluidic device
Microscope
Microscope electron
Stain
Sterile
Corresponding Organization : Syracuse University
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Variable analysis
independent variables
- Microfluidic device used to obtain aggregated proteins
- Aggregated proteins collected
- Negative stain EM data collected
- Cryo-EM data collected
- Ultrafiltered deionized water used to flush surfaces
- Remaining liquid in reservoirs refrigerated along with collected liquid
- Procedures for collecting aggregated proteins from shaking experiments and EM sample preparation described previously
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