INSIDIA
If you want to do spheroid segmentation, click "No, I want to do spheroid segmentation" in this menu.
The first grayscale image will be opened and an attempt to automatically retrieve spheroid mass will be done.
If you're working with fluorescence images please check the box in the menu
INSIDIA will display the original grayscale image with the spheroid mass outlined in yellow and a MENU entitled Spheroid Segmentation. In this menu, you can accept automatic threshold and proceed to next image. If you are not satisfied by this automatic step you can perform manual thresholding or Frangi filtering.
If you select Frangi filter, you should provide parameters of number, maximum and minimum (see http://imagej.net/Frangi for the meaning of each parameter). INSIDIA will calculate again the spheroid outline and you can accept or repeat this operation.
If you select manual thresholding a MENU entitled ManualThreshold will appear together with the Threshold Menu of ImageJ. In this case, the grayscale image will become blue in some areas. Those areas will be considered as spheroid mass. You can define manually blue regions by using the slide bars in the Threshold Menu and change the cut-offs of the histogram. Once satisfied with the blue area considered, click OK in the ManualThreshold menu. INSIDIA will calculate again the spheroid outline and you can accept or repeat this operation.