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Krios-K2 Presets used for 2.8 A proteasome reconstruction at TSRI¶
Tecnai Krios 300 kV high tension, Gun Lens 5 extraction voltage 4300, 70 um C2 aperture and 100 um objective aperture (except when obtaining grid atlas).
Gatan camera dimension 3710(w)x 3838(h) after rotation to Leginon standard. The size of the beam on the scope main viewing screen always covers the 1.2 um hole at en preset on our scope.
Pixel size is 1.21 A at 22500x scope nominal mag.:
| Magnification: | Preset name: | Image Shift (x,y): | Dimension: | Binning: | Camera Dose (e/camera pixel/s): | Exposure Time (ms): | Specimen Dose Rate (e/A^2): | Spot Size: | Defocus (m): | C2 (um): | frame saving: |
|---|---|---|---|---|---|---|---|---|---|---|---|
| 81 | gr | Aligned | 927x959 | 4 | 8 | 100 | n/a | 10 | 0.0 | 70 | no |
| 165 | sq | Aligned | 927x959 | 4 | 8 | 100 | n/a | 9 | -2e-4 | 70 | no |
| 1700 | hl | Aligned | 927x959 | 4 | 8 | 300 | 0.008 | 9 | -1.5e-4 | 70 | no |
| 22500 | fc | 0,0 | 924x924 | 2 | >=8* | 500 | 8.8 | 6 | -2.9e-7 | 70 | no |
| 22500 | fm | 0,0 | n/a | n/a | high | n/a | high | 5 | n/a | 70 | no |
| 22500 | fa | 0,0 | 927x959 | 4 | >=8* | 200 | 3.5 | 6 | -1.5e-6 | 70 | no |
| 22500 | en | 0,0 | 7420x7676 | 1 | 8 e/physical pixel/s and ~1.9 um on the specimen | 7600 | 36 | 8 | (-1e-6 to -2e-6) | 70 | 200 ms/frame |
- Note: This experiment has an additional preset fm used for melting the ice exclusively. The screen goes down during the melt, therefore, it does not cause radiation damage on the camera. Do not attempt to acquire an image with this preset.
- Note: If the correlation or power spectrum don't work well enough for the standard Camera Dose Rate of 10 e/pixel/s for fc and fa, you may increase it more so not to have to use longer exposure time. The recommended value in literature is meant to give highest DQE at all sampling frequencies. Using a higher camera dose by 50% will certainly not damage the sensor.
- Note: no defocal pair was acquired in this experiment. Therefore, no ef preset.
Updated by Anchi Cheng over 7 years ago · 4 revisions