80 Chapter 3 It should be noted that multiple compute requests can be simultaneously handled. The size of the CZI file should be smaller than 1.5GB. The width and height of the stack images can be in any size. However, the bigger the image size is the more computational time would be needed. For each request separately, all above-mentioned computations will be computed and represented in the redirected page within a maximum of 12 hours after uploading the CZI file. Currently, the results will be deleted after a week of the submitted request. VALIDATION AND RESULTS To validate the accuracy of OrganelX along with its interactive tool we defined and compared two scenarios. Segmentation and area calculation for one stack image For a randomly chosen stack image, we manually annotated 95 organoids and compared their cross-sectional area against the computed area by OrganelX. It is to be mentioned that the segmentation of one stack image using OrganelX was completed in a few minutes while the time for the manual segmentation for the same stack was around 1h. Furthermore, while manual counting allowed us to segment 95 organoids, OrganelX was able to count twice as many organoids. Figure 2A shows the chosen stack image in addition to the manually segmented organoids in red. The black and blue points in Figure 2B denote the cross-sectional area of those manuallly and OrganelX segmented organoids, respectively. The results demonstrate that the overall computed areas are nearly identical with a minor difference for some organoids. To demonstrate the accuracy of this result in a number, the average of the absolute differences between manual and OrganelX segmented area was calculated, and divided by the total manual segmented area. An inaccuracy of less than 0.001% was obtained.
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