Scintillate: An open-source graphical viewer for time-series calcium imaging evaluation and pre-processing.
Calcium imaging is based on the detection of minute signal changes in an image time-series encompassing pre- and post-stimuli. Depending on the function of the elicited response, change may be pronounced, as in the case of a genetically encoded calcium-reporter protein, or subtle, as is the case in a bath-applied dye system. Large datasets are thus often acquired and appraised only during post-processing where specific Regions of Interest (ROIs) are examined.



The scintillate software provides a platform allowing for near instantaneous viewing of time-sequenced tiffs within a discrete GUI environment. Whole sequences may be evaluated. In its simplest form scintillate provides change in florescence (ΔF) across the entire tiff image matrix. Evaluating image intensity level differences across the whole image allows the user to rapidly establish the value of the preparation, without a priori ROI-selection. Additionally, an implementation of Independent Component Analysis (ICA) provides additional rapid insights into areas of signal change.



We imaged transgenic flies expressing Calcium-sensitive reporter proteins within projection neurons and moth mushroom bodies stained with a Ca2+sensitive bath-applied dye. Instantaneous pre-stimulation background subtraction allowed us to appraise strong genetically encoded neuronal Ca2+responses in flies and weaker, less apparent, responses within moth mushroom bodies.



At the time of acquisition, whole matrix ΔF analysis alongside ICA is ordinarily not performed. We found it invaluable, minimising time spent with unresponsive samples, and assisting in optimisation of subsequent acquisitions.



We provide a multi-platform open-source system to evaluate time-series images.
Division of Chemical Ecology, Department of Plant Protection Biology, Swedish University of Agricultural Sciences, P.O. Box 102, SE-230 53, Alnarp, Sweden. Electronic address: ian.dublon@slu.se.
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IAN
Dublon
Lund University Bioimaging Center, Faculty of Medicine, Lund University, Lund, Sweden.
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M
Nilsson
Division of Chemical Ecology, Department of Plant Protection Biology, Swedish University of Agricultural Sciences, P.O. Box 102, SE-230 53, Alnarp, Sweden.
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A
Balkenius
Division of Chemical Ecology, Department of Plant Protection Biology, Swedish University of Agricultural Sciences, P.O. Box 102, SE-230 53, Alnarp, Sweden.
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P
Anderson
Division of Chemical Ecology, Department of Plant Protection Biology, Swedish University of Agricultural Sciences, P.O. Box 102, SE-230 53, Alnarp, Sweden.
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MC
Larsson
D000818:Animals; D030801:Animals, Genetically Modified; D002118:Calcium / Q000378:metabolism*; D004331:Drosophila melanogaster; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007091:Image Processing, Computer-Assisted; D052139:Intracellular Calcium-Sensing Proteins / Q000235:genetics / Q000378:metabolism; D018402:Manduca; D024521:Mushroom Bodies / Q000166:cytology / Q000378:metabolism; D009474:Neurons / Q000378:metabolism*; D061848:Optical Imaging; D025341:Principal Component Analysis; D012984:Software; D013997:Time Factors
D052139:Intracellular Calcium-Sensing Proteins; D049452:Green Fluorescent Proteins; D002118:Calcium
Calcium imaging; Fluorescence; GUI; ICA; Pre-processing; optical imaging