Extract Templates Directly From Micrograph


  1. Define a new project space with the name "KLH".
  2. Load micrograph "KLH_02_01.mrc".
  3. Quality inspection: intensity histogram seems okay.
  4. Quality inspection: activate SPEC button, inspect spectra of selected regions.
  5. Filter the image to reduce noise and enhance contrast. In Image Pre-processing box, set Low-pass to 140, and High-pass to 1000. Click the Filter button.
  6. The templates will come from the micrograph directly. Click the Filament button at the bottom of the window. A rectangular monitor will show up in the panel.
  7. Set particle diameter (Size) to 800 in the Filament Parameters box.
  8. Identify a clean top-view, draw a box around the particle (press CTRL, left-click and drag). Set the scaling of the monitor to 2 or 3 in order to display the full particle. Save the image (Save button) as "topView.mrc" (see example).
  9. Identify a clean side-view, draw a box around the particle, and save the image as "sideView.mrc" (see example).
  10. Go to Menu : Function : Image Stack Editor, click New and load "topView.mrc". Then, click Add to append "sideView.mrc". Crop both images to an appropriate size (200-pixels in the example) using the Crop function in the Image Processing box. Click Save to save the templates as "klhTemp.mrc" (see example).
  11. Load template image "klhTemp.mrc" (Menu : Data : Template : Load MRC).
  12. Erase the default template masking by clicking Clear in Manual Masking box.
  13. Set Frame to 1 in Manual Masking box, then click Edit. The top-view is displayed. Activate the circle-drawing button on the toolbar (mouse click or press F3 key). Position the circle around the top-view at a proper radius (Left-click-drag for shifting, Right-click-drag for radius resizing). Deactivate the circle-drawing button, a circular mask will be drawn. Release the Edit button.
  14. Set Frame to 2 in Manual Masking box, then click Edit. The side-view is displayed. Activate the line-drawing button on the tool bar (mouse click or press F4 key). Set the brush-size to 16 and draw a rectangular mask around the particle. Deactivate the line-drawing button, release the Edit button (see example).
  15. Micrograph screening can now start following the normal procedure.