Measurement of intracellular calcium gradients in single living cells using optical sectioning microscopy

Rao V. Yelamarty; Joseph Y. Cheung

doi:10.1117/12.59540

26 June 1992 Measurement of intracellular calcium gradients in single living cells using optical sectioning microscopy

Rao V. Yelamarty, Joseph Y. Cheung

Proceedings Volume 1660, Biomedical Image Processing and Three-Dimensional Microscopy; (1992) https://doi.org/10.1117/12.59540
Event: SPIE/IS&T 1992 Symposium on Electronic Imaging: Science and Technology, 1992, San Jose, CA, United States

Abstract

Intracellular free calcium has been recognized as a regulator of many cellular processes and plays a key role in mediating actions of many drugs. To elucidate subcellular spatial calcium changes throughout the cell in three dimensions (3-D), optical sectioning microscopy was applied using digital imaging coupled fluorescence microscopy. The cell was loaded with a fluorescent indicator, fura-2, and a stack of sectional fluorescent images were acquired, digitized and finally stored on-line for post image analysis. Each sectional image was then deconvolved, to remove contaminating light signals from adjacent planes, using the Nearest Neighboring Deconvolution Algorithm (NNDA) and the overall imaging system's empirical Point Spread Function (PSF) that is measured with a 0.25 micrometers fluorescent bead. Using this technique, we measured that the addition of growth factors caused a 2 - 3 fold increase (1) in nuclear calcium compared to cytosolic calcium in blood cells and (2) in both nuclear and cytosolic calcium in liver cells. Such spatial information, which is important in understanding subcellular processes, would not be possible to measure with other methods.

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Rao V. Yelamarty and Joseph Y. Cheung "Measurement of intracellular calcium gradients in single living cells using optical sectioning microscopy", Proc. SPIE 1660, Biomedical Image Processing and Three-Dimensional Microscopy, (26 June 1992); https://doi.org/10.1117/12.59540

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