High-Resolution Calcium Imaging Method for Local Calcium Signaling
All eukaryotic cells respond to extracellular signals in a physiologically meaningful way. For multicellular organisms, physiologically relevant cooperation is only possible, if cell-cell communication works properly. Here, the extracellular signals must be translated into intracellular signals that ultimately result in cellular responses. This process is termed signal transduction or signaling. Ca2+ signaling has been developed in almost all eukaryotic cells. The cellular components used for this highly versatile signaling system are often termed “Ca2+ toolbox”. Besides Ca2+ pumps and Ca2+-binding proteins, the Ca2+ channels that are located in the plasma membrane and intracellular membranes and the Ca2+-mobilizing second messengers are major players in shaping the four-dimensional nature of Ca2+ signals.
Here, we report on methodological developments to acquire and analyze cellular Ca2+ signals with high temporal and spatial resolution with specific focus on (1) photobleaching of Ca2+ indicators at high acquisition rate, (2) determination of system noise and spatiotemporal detection limits, and (3) image processing.
Key wordsCa2+ signaling Ca2+ microdomain cell signaling High-resolution imaging Live cell imaging Deconvolution
The authors acknowledge financial support by Deutsche Forschungsgemeinschaft (SFB1328 projects A01 (to AHG), A02 (to IMAW and RW), grants GU 360/15-1 and 15-2 (AHG)), Joachim-Herz-Stiftung, Infectophysics Consortium, project 04, and Landesforschungsförderung Hamburg ReAd Me (project 1, IMAW and AHG).
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