SparkMaster: automated calcium spark analysis with ImageJ
SparkMaster: automated calcium spark analysis with ImageJ
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DOI：
10.1152/ajpcell.00586.2006
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发表时间：
2007-09-01
影响因子：
5.5
通讯作者：
Bers, Donald M.
中科院分区：
文献类型：
--
作者：
Picht, Eckard;Zima, Aleksey V.;Bers, Donald M.
钙火花是细胞内钙储存的基本钙释放事件，几乎在所有类型的肌肉中都可以观察到。通常情况下，钙火花测量线扫描模式与共焦激光扫描显微镜，产生二维图像（距离与时间）。对这些图像的手动分析是耗时的，并且容易出错以及调查者偏见。因此，我们开发了SparkMaster，这是一种自动分析程序，可以进行快速可靠的火花分析。基础分析算法改编自Cheng等人开发的火花分析的基于阈值的标准方法（Biophys J 76：606-617，1999），并且在此在可免费获得的图像处理软件ImageJ中实现。SparkMaster提供了一个图形用户界面，通过该界面可以选择所有分析参数和输出选项。分析包括一般图像参数（检测到的火花的数量、火花频率）和单个火花参数（振幅、半最大振幅处的全宽度、半最大振幅处的全持续时间、全宽度、全持续时间、到达峰值的时间、火花上升冲程的最大陡度、火花衰减的时间常数）。我们使用具有嵌入到具有不同信噪比的背景中的合成火花的图像来验证该算法，以确定高灵敏度与低频率的假阳性检测相结合的分析标准。最后，我们应用SparkMaster来分析在完整和透化的心室心肌细胞、透化的哺乳动物骨骼肌和完整的平滑肌细胞中测量的火花的实验数据。我们发现SparkMaster提供了一种可靠，易于使用和快速的方法来分析各种实验条件下的钙火花。
Ca sparks are elementary Ca-release events from intracellular Ca stores that are observed in virtually all types of muscle. Typically, Ca sparks are measured in the line-scan mode with confocal laser-scanning microscopes, yielding two-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 that 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 76: 606-617, 1999) 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) and individual spark parameters (amplitude, full width at half-maximum amplitude, full duration at half-maximum amplitude, 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.