Q: “Immunohistochemistry was performed on my lung biopsies and now I have to analyse my staining. I would like to get a value for the colour intensity of my staining. … In the output of Pixcavator a mean value for grayness is given, but I don’t think all my brown staining is measured. Even if I adjust the threshold for size, not all objects are captured with a red or green contour.  ”

1. The difference in gray isn’t more pronounced because what is displayed is the average not the weighted average. So, one large dark object is outweighed by smaller, lighter ones. If you hover over objects (or look at the table) one at a time, you see a more noticeable difference. You can also save the data as a spreadsheet and compute the weighted averages. Certainly, if the difference is still too small, contrast enhancement would help.

2. It’s better to choose the green channel (or blue) than red. The features are much more distinct. If you click on “Display channel”, you’ll see the difference.

3. You are interested in dark objects only, red contours. So, the light ones only skew the averages. Click “Unmark light”.

4. The “Size” slider does not seem to reveal the features quite well. I used the simple thresholding instead, i.e., the “Intensity, dark” slider.

A screenshots is below. (This example may be somewhat similar: Measuring staining in the liver.)

More analysis here… 

Other examples of image analysis