Connectivity assay workflow

Purpose

The connectivity assay is a second order probe used to makes estimates about the total number of connections or trabeculae.

More information on connectivity in stereology is available at stereology.info.

We recommended that you place a reference point in an easy to find and recognizable location before starting this probe workflow. When working with a live camera feed, the placement of the reference point is essential for maintaining accurate alignment between a data file and its corresponding experimental sample when collecting data over multiple sessions. If a reference point has not been placed before opening a workflow, Stereo Investigator will automatically place it in the top left corner of the Main window. This location may not be ideal for resuming data collection in subsequent sessions.

Starting the connectivity assay workflow

  1. Click the Connectivity assay workflow button on the Probes ribbon.

    If you haven't used the workflow recently, find the Connectivity assay workflow button in the Second order drop-down menu in the All Probes section of the Probes ribbon.

  2. In the dialog box that opens, the following choices may be available, depending on what files, if any are currently open. Choose what you would like to do:

    • Start a new subject: Count objects on a live-camera feed from your microscope or in an image file, containing the entire region to be counted that you have already opened.
    • Load subject data from existing file: Continue working in a stored data file that is not currently open.

Workflow overview

The workflow is dynamic; your choices determine which steps are included, and what information is requested and displayed. Because of this, the step numbers you see onscreen in Stereo Investigator may be different than those in the complete workflow described here.

Click any of the underlined steps to jump to the instructions:

Indicate the region(s) of interest

1. Set up the subject

2. Set microscope to low magnification

3. Trace your region(s) of interest

4. Set microscope to high magnification

Define probe configuration

5. Define the counting frame size

6. Define SRS grid layout

7. Save sampling parameters

Align and Mark the Tissue

8. Acquire alignment images

9. Match tissue sections

10. Mark tissue

11. View the results

Commands available in every step of the workflow:

New workflow: Click the new workflow button to start over; the settings will revert to the defaults or those specified in the previous completed workflow.
Previous step / Next Step: Click to advance in the workflow or revisit a previous step. Alternatively, you can click the steps listed at the top of the workflow to jump to that step.
Click the help link to view information in the User Guide on completing the current step in the workflow.

All stereological formulas