Gradual loss of fluorescence during the capturing of time-lapse sequence of fluorescently labeled samples is known as photobleaching and is a general problem in biological imaging. The bleaching could be avoided by tuning the microscopy set-ups, but when such adjustments only partially works image sequence with decaying fluorescence intensity should be corrected in order to deploy precise segmentation or for the quantification of intensity dynamics. We implemented an ImageJ plugin that allows the user to compensate the photobleaching artifact to estimate the non-bleaching condition, with choice of three different algorithms: simple ratio, exponential fitting and histogram matching methods. In the presentation, details and characteristics of each algorithm will be expained based on application to actual image sequences.
Photobleaching, 3D, histogram matching, curve fitting, time-lapse
I work in EMBL Heidelberg as an image analysis specialist and collaborate mainly with cell and developmental biologist. My research interests are: diffusion in cytoplasm, intracellular vesicle transport, cell migration mechanics, coordination of multicellular movement and taxis.
Presenting author: Kota Miura
Organisation: EMBL Heidelberg