For a successful Guided Acquisition experiment, you need to prepare an overview and a detailed experiment as well as an image analysis setting. If you want to process your overview image before it is analyzed, you also need a suitable setting for each processing step or function you want to execute.

Overview scan

You have defined the experiment for the overview scan. Typically, the overview scan is using a lower magnification in combination with a tile experiment.

Processing setting(s)

You have defined a suitable setting for each processing step or function you want to execute. For more information, see General Settings.
Note that if you want to use Shading Correction with a reference image, you have to define your setting in Batch mode!

Image analysis setting

You have defined a suitable image analysis setting, see Creating a New Image Analysis Setting.

Detailed scan

You can perform for example the following experiments:

• simple Z-stacks with a high-NA objective lens
• Multi-Channel Z-stacks using an optical sectioning method like SD, Apotome or Airyscan
• Tile experiments
  Note: If you have activated Auto-Distribute for New Tile Regions for support point distribution in the Tiles tool, this is saved in the experiment and used for Guided Acquisition as well.
• Time Series
  Note that for detailed experiments that are defined as a time series, a time series for each detected object is performed before moving to the next detected object. In order to create a time series which acquires the first time point for all detected objects before starting the acquisition for the second time point, use the Repeat Detailed Scan option.

Parcentricity

If you use different detectors for the overview and the detailed scan, it might be necessary to correct for the shift between both detectors (to ensure parcentricity). For this you have to take an image at the exact same position with both cameras and then determine the offset between the two (to ensure parcentricity). The reference to calculate this offset is the image taken with the camera for the overview experiment. You can then enter the values for the shift in X and Y in the guided acquisition setup.

Focus strategies

You have several options, to perform a focus strategy.

Detailed scan and overview scan can be defined with their own focus strategy using Focus Surface and/or Software Autofocus.

During the Guided Acquisition experiment, you can define additional focusing steps. They are independent from the focus strategy defined on the Acquisition tab, see Focus Strategy Tool.

For more information on Guided Acquisition, see Performing a Guided Acquisition.