SRS Image Series workflow > Acquire >

9. Pre-focus

Method for identifying top of section

Choose one of the following:

  • At every ... acquisition site: This is the recommended method.

    • Type in the number (N) to indicate that you will identify the top of the section at every "N" acquisition sites.

    • (optional) Select random ...sites: if you chose to identify the top of the section at fewer than every acquisition site (i.e., you entered a number >1 in the box for At every ... acquisition site), you can check the box to have Stereo Investigator automatically select the focus sites.

    • Click Update Display to see your selected focus sites

  • Interactive site selection: Choose focus sites manually. This may be of interest if you know that there are differences in the specimen thickness and you know where they occur.

    • Click within contours to manually select focus sites.
    • To deselect a focus site, click within the contour again.

  • At current Z position: Acquire each stack in the series at the current Z position at each site.

    This option is recommended only for very consistent, flat tissue that has little or no variation in Z-step.

 

Set Top of Section button

If you chose At every ... acquisition site or Interactive site selection as the method for identifying top of section, you must establish the top of the specimen at several sites (the number of sites depends on the options chosen).

  1. Click Set top of section. The stage moves to each focus site and the Focus Site dialog box opens.
  2. Adjust the focus to the top of the desired focal plane.

  3. Click Next to move to the next site.

    A running count of completed focus sites is displayed at the bottom of the screen.

  4. Repeat until all sites have been visited.

Buttons in the Focus Site dialog box

  • Previous: Revisit the previous focus site.
  • Skip: Skip the current focus site.
  • Next: Move to the next site.

  • More...: Opens the Device Control window that provides easy access to:

Surface Map

This section appears once you have identified the top of section for each focus site; it is not displayed if you chose At current Z position as the method for identifying the top of section.

  • In the Main window, a grid of your focus sites, with their Z-positions in red is displayed over the contour(s).
  • Show/Hide surface map: Click to show or hide a surface map designed to help you assess the topology of the specimen and to determine if additional focus sites are needed to provide more clarity in the image.