Combined system for high time resolution dual excitation fluorescence photometry and fluorescence imaging of calcium transients in single normal and diseased skeletal muscle fibers

Dietmar Uttenweiler; Reinhold Wojciechowski; Makoto Makabe; Claudia Veigel; Rainer H.A. Fink

doi:10.1117/12.197514

22 December 1994 Combined system for high-time-resolution dual-excitation fluorescence photometry and fluorescence imaging of calcium transients in single normal and diseased skeletal muscle fibers

Dietmar Uttenweiler, Reinhold Wojciechowski, Makoto Makabe, Claudia Veigel, Rainer H.A. Fink

Author Affiliations +

Proceedings Volume 2328, Biomedical Optoelectronic Devices and Systems II; (1994) https://doi.org/10.1117/12.197514
Event: International Symposium on Biomedical Optics Europe '94, 1994, Lille, France

Abstract

Fast photometric measurements and video-imaging of fluorescent indicators both are powerful tools in measuring the intracellular free calcium concentration of muscle and many other cells. as photometric systems yield a high temporal resolution, calcium imaging systems have high spatial but significantly reduced temporal resolution. Therefore we have developed an integrated system combining both methods and based mostly on standard components. As a common, sensitive Ca²⁺- indicator we used the fluorescent probe Fura-2, which is alternatingly excited for ratio measurements at 340/380 nm. We used a commercially available dual excitation photometric system (OSP-3; Olympus) for attaching a CCD-camera and a frame grabber board. To achieve the synchronization we had to design circuitries for external triggering, synchronization and accurate control of the filter changer, which we added to the system. Additionally, the software for a triggered image acquisition was developed. With this integrated setup one can easily switch between the fast photometric mode (ratio frequency 100 Hz) and the imaging mode (ratio frequency 4.17 Hz). The calcium images are correlated with the 25 times faster spot measurements and are analyzed by means of image processing. With this combined system we study release and uptake of calcium ions of normal and diseased skeletal muscle from mdx mice. Such a system will also be important for other cellular studies in which fluorescence indicators are used to monitor similar time dependent alterations as well as changes in cellular distributions of calcium.

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Dietmar Uttenweiler, Reinhold Wojciechowski, Makoto Makabe, Claudia Veigel, and Rainer H.A. Fink "Combined system for high-time-resolution dual-excitation fluorescence photometry and fluorescence imaging of calcium transients in single normal and diseased skeletal muscle fibers", Proc. SPIE 2328, Biomedical Optoelectronic Devices and Systems II, (22 December 1994); https://doi.org/10.1117/12.197514

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