BIO407 Group6 Image processing of live cells, treated with DFX

## Study: This data is part of the practical course BIO407 2025 at the University of Zurich, titled 'Advanced Microscopy: From preparation to data and visualization'. The effects of Deferasirox (DFX) on mitochondria morphology were examined using different microscopy techniques. The data is originally from the paper: Gottwald EM, Schuh CD, Dr點ker P, Haenni D, Pearson A, Ghazi S, Bugarski M, Polesel M, Duss M, Landau EM, Kaech A, Ziegler U, Lundby AKM, Lundby C, Dittrich PS, Hall AM. The iron chelator Deferasirox causes severe mitochondrial swelling without depolarization due to a specific effect on inner membrane permeability. Sci Rep. 2020 Jan 31;10(1):1577. doi: 10.1038/s41598-020-58386-9 . PMID: 32005861 ; PMCID: PMC6994599. ## Study Component: Job: Image Processing Task: 4) Timelapse of mitochondria in deferasirox (DFX) treated cells Timelapse data was analyzed: Mitochondria were segmented, their morphology and signal intensity were measured over time, and plotted. ## Biosample: Opossum kidney (OK) cells (kind gift from the group of Prof O. Devuyst (Physiology, University of Zurich)) ## Specimen: DFX treatment: 200uM Dyes: -Mitochondria-GFP BacMam 2.0 -TMRM (mitochondrial membrane potential dependent dye) ## Image Acquisition: Images were acquired using a Leica SP8 inverse STED 3x. Acquired channels: Channel 1: Mitochondria-GFP, 488nm excitation, 493nm-548nm emission Channel 2: TMRM, 553nm excitation, 564nm-650nm emission Objective: HC PL APO CS2 100x/1.40 OIL ## Image Data: Raw timelapse data: 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression.lif Segmentation masks of mitochondria of channel with "Calculate Threshold for each image": C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_channel1_gaussian_otsu_open_watershed.tif Mean GFP signal and circularity of segmented mitochondria at each timepoint channel 1: Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1_channel1.csv Mean GFP signal circularity of segmented mitochondria at each timepoint channel 2: Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1_channel2.csv Plot of mean GFP signal in mitochondria over time of channel 1: Plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1pixel_channel1.png Plot of mean GFP signal in mitochondria over time of channel 2: Plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1pixel_channel2.png Plot of mean circularity in mitochondria over time of channel 1: Plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_circularity_channel1.png ## Image Correlation C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_channel1_gaussian_otsu_open_watershed.tif contains the segmentation masks for the timelapse in 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression.lif Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1_channel1.csv contains the measurements for the timelapse in C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_channel1_gaussian_otsu_open_watershed.tif redirected to channel 1 Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1_channel2.csv contains the measurements for the timelapse in C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_channel1_gaussian_otsu_open_watershed.tif redirected to channel 2 Plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1pixel_channel1.png contains the mean intensity plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1_channel1.csv Plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1pixel_channel2.png contains the mean intensity plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1_channel2.csv Plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_circularity_channel1.png contains the mean circularity plot of Summary of C1-151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression-1_gaussian_otsu_open_watershed_MIN1_channel1.csv ## Image Analysis We used Fiji to extract channel 1 of our raw timelapse image. We then applied Gaussian Blur, Otsu Thresholding ("Treshold for each image" + "Auto"), Binary-->Open and Watershed to get a binary mask of our raw timelapsed image. Circularity and mean intensity were then measured by using the mask of channel 1 and redirected to the raw timelapse file for channel 1 and channel 2.