Physical Fractionator workflow
Overview
Physical Fractionator uses a two-stage systematic sampling method to estimate the number of objects in a specified region of an organ. This method combines the Disector method with the fractionator sampling method.
Like Optical Fractionator, Physical Fractionator is typically used when the population is too large to count exhaustively. Since it relies on the Disector method, pairs of thin sections are required.
About the Disector Method
The disector is a 3D stereological probe with one unique and very important attribute: it samples objects with a probability that is proportional to their number, not their size. Because of this, the disector can be used to count objects without having to make any assumptions about their size, shape, or orientation. In its simplest form, the disector involves the use of two sections, hence the name disector.
The counting rule used with the disector probe is simple. One counts the objects that are present on the second section of a pair of adjacent sections that are not present on the first, as one proceeds systematically through a series of sections that includes the region under consideration. An equally valid counting rule is to count objects that are observable in the first section and not in the second. Essentially what one is doing is determining whether or not a unique point on each object, the "top," or as in the second case, the "trailing edge," lies within the volume sampled by the disector.
Starting the workflow
- Click Probes>Number>Mono-layer>Physical Fractionator workflow.
- You are prompted to start new subject (Start a new subject) or to continue with a subject (Load subject data from existing file).
If you select Load subject data from existing file:
- Select the data file that has already been started by clicking on the Browse button.
- You are directed to step 10 - Mark cells.
Workflow steps
Indicate the Region(s) of Interest
- Step 1: Set up the subject
- Step 2: Set system to low magnification
- Step 3: Trace your region(s) of interest
- Step 4: Set system to high magnification
Define Probe Configuration
- Step 5: Define the counting frame size
- Step 6: Define SRS grid layout
- Step 7: Save sampling parameters
Align and Mark the Cells
- Step 8: Acquire alignment images
- Step 9: Match tissue sections
- Step 10: Mark cells
- Step 11: View the results