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Am J Physiol Cell Physiol (March 21, 2007). doi:10.1152/ajpcell.00586.2006
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Submitted on November 21, 2006
Accepted on March 20, 2007

SparkMaster - Automated Calcium Spark Analysis with ImageJ

Eckard Picht1, Aleksey V Zima1, Lothar A Blatter1, and Donald M. Bers1*

1 Department of Physiology, Loyola University Chicago, Maywood, Illinois, United States

* To whom correspondence should be addressed. E-mail: dbers{at}lumc.edu.

Calcium (Ca) sparks are elementary Ca release events from intracellular Ca stores which are observed in virtually all types of muscle. Typically, Ca sparks are measured in the linescan mode with confocal laser scanning microscopes, yielding 2-dimensional images (distance vs. time). The manual analysis of these images is time consuming and prone to errors as well as investigator bias. Therefore, we developed SparkMaster, an automated analysis program which allows rapid and reliable spark analysis. The underlying analysis algorithm is adapted from the threshold-based standard method of spark analysis developed by Cheng et al. (Biophys J 1999;76:606-17) and is implemented here in the freely available image processing software ImageJ. SparkMaster offers a graphical user interface through which all analysis parameters and output options are selected. The analysis includes general image parameters (number of detected sparks, spark frequency) as well as individual spark parameters (Amplitude, FWHM, FDHM, Full Width, Full Duration, Time-to-Peak, maximum steepness of spark upstroke, time constant of spark decay). We validated the algorithm using images with synthetic sparks embedded into backgrounds with different signal-to-noise ratios to determine an analysis criteria at which a high sensitivity is combined with a low frequency of false positive detections. Finally, we applied SparkMaster to analyze experimental data of sparks measured in intact and permeabilized ventricular cardiomyocytes, permeabilized mammalian skeletal muscle, and intact smooth muscle cells. We found that SparkMaster provides a reliable, easy to use and fast way of analyzing Ca sparks in a wide variety of experimental conditions.




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