2.3 | Image Processing
Image processing techniques were utilized to retrieve raw intensity profiles from stacks of images for each respective treatment and their duplicates during the 8-hour period of time-lapse fluorescence microscopy. Raw red channel image stacks are processed using ImageJ (National Institutes of Health, Bethesda, MD). First, contrast enhancement is applied to obtain mitochondrial contours. This allows for a ROI (region of interest) to be selected and applied over all 24 images in a stack. For quantification, the selected ROI is used to retrieve mitochondrial fluorescent intensity presented as A.F.U (arbitrary fluorescent units) and background (dark regions) for background subtraction. Intensity profiles are fitted to sigmoidal (Equation 1) and linear (Equation 2) models using MATLAB R2022a (MathWorks Inc., Natick, MA) to visualize trends in drug treatments.
\(y=\frac{1}{1+e^{-x}}\) (1)
\(y=mx+b\) (2)