ScintillateAn open-source graphical viewer for time-series calcium imaging evaluation and preprocessing Dublon, I. A N; Nilsson, Markus; Balkenius, A.; Anderson, P.; Larsson, M. C. Methods, 273, 120-127. DOI: 10.1016/j.jneumeth.2016 General rights Copyright and moral rights for the publications made accessible in the public portal are retained by the authors and/or other copyright owners and it is a condition of accessing publications that users recognise and abide by the legal requirements associated with these rights.• Users may download and print one copy of any publication from the public portal for the purpose of private study or research.• You may not further distribute the material or use it for any profit-making activity or commercial gain • You may freely distribute the URL identifying the publication in the public portal
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Abstract BackgroundCalcium 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 calciumreporter 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.
New methodThe 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.
ResultsWe imaged transgenic flies expressing Calcium-sensitive reporter proteins within projection neurons and moth mushroom bodies stained with a Ca 2+ sensitive bath-applied dye. Instantaneous pre-stimulation background subtraction allowed us to appraise strong genetically encoded neuronal Ca 2+ responses in flies and weaker, less apparent, responses within moth mushroom bodies.
Comparison with existing methodsAt 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.
ConclusionsWe provide a multi-platform open-source system to evaluate time-series images.Abbreviations: ΔF, change of intensity between frames; ICA, independent component analysis