Our objective is to count (in histologic images) both mast cells (our identified primary object) as well as the number of granules surrounding each mast cell (within a specified perimeter). We have been able to successfully count and identify the number of mast cells in an image, however we have so far been having difficulty identifying and counting the small sattelite granules within a specified perimeter surrounding each mast cell. We have tried using the Measure Object Neighbors module within Cell Profiler unsuccessfully so far - the module actually identifies other PRIMARY OBJECTS (in this case, other mast cells) and labels them as neighbors. We would like the software to identify and count the small, sattelite granules surrounding each mast cell as NEIGHBORS (instead of other mast cells). Is there either a module available or a way for us to adapt an already existing module in order for us to define an area/perimeter around an identified primary object (i.e. mast cell) and count much smaller neighbors (i.e. small, sattelite granules) within this area/perimeter?
This is a great question, and yes, CellProfiler should be able to accomplish this as is.
(1) Use IdentifyPrimAutomatic to define the mast cells (only) as you have been
(2) Use IdentifySecondary to define a region around and including each mast cell. This can be done simply as a constant width (Distance-N option) or using other image information (Propagation, Distance-B, or Watershed methods) depending on your stains.
(3) Use IdentifyTertiarySubregion to define the region surrounding the mast cells, excluding the mast cells themselves
(4) Use an additional IdentifyPrimAutomatic module to define only the “small satellite granules”. Hopefully setting the “Typical diameter of objects” to a low value will define these as a separate population from the mast cells. If not, you may need to use a Measurement module with a FilterByObjectMeasurement module on whatever Measurement will filter out the mast cells.
(5) Here’s the key step: Use the Relate Module to relate the children (granules) to their parents (the regions around the mast cells from IDTertiary). You might also relate the IDSecondary objects to the granules with another Relate module in case some granules fall within the defined mast cells.
(6) ExportToExcel and choose all your relevant objects. Look for the Parent/Children columns in the granules/tertiary regions, respectively.
Hope this helps!