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  A confocal scanning laser microscope for quantitative ratiometric 3D measurements of [Ca2+] and Ca2+ diffusions in living cells stained with Fura-2

Helm, P. J., Franksson, O., & Carlsson, K. (1995). A confocal scanning laser microscope for quantitative ratiometric 3D measurements of [Ca2+] and Ca2+ diffusions in living cells stained with Fura-2. Pflügers Archiv: European Journal of Physiology, 429(5), 672-681. doi: 10.1007/BF00373988.

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Alternative Title : A confocal scanning laser microscope for quantitative ratiometric 3D measurements of [Ca2+] and Ca2+ diffusions in living cells stained with Fura-2

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EurJPhysiol_429_1995_672.pdf (Any fulltext), 2MB
 
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 Creators:
Helm, Paul Johannes1, Author              
Franksson, Olof, Author
Carlsson, Kjell, Author
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1Department of Cell Physiology, Max Planck Institute for Medical Research, Max Planck Society, ou_1497701              

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 Abstract: A confocal scanning laser microscope (CSLM) for observation and quantitative ratiometric measurements of the intracellular dynamics of Ca2+ ions in living neurons has been developed. The instrument consists of a UV-enhanced CSLM, an optical arrangement providing simultaneous excitation at two wavelengths, an electronic arrangement for processing the simultaneous fluorescence response, and software for computing the absolute Ca2+ concentrations, ([Ca2+]). The instrument can be used for any excitation ratiometric measurements, provided that the dye substance used is excitable by wavelengths between 334 nm and 750 nm (such as, e.g. Fura-2). The spatial resolution of the CSLM, as well as a temporal resolution of 20 ms per line (maximum sampling rate) for dynamic measurements are provided by the instrument. Using Fura-2 in calibrated Ca2+ buffer solutions, the instrument measures [Ca2+] between 0 and 1.35 mumol.l-1 with an error of less than 1%. The capability of the instrument to measure absolute [Ca2+] was verified by recording fluorescence images of test solutions with well defined [Ca2+] values (Molecular Probes, Eugene, Ore., USA, C-3009 calibration solutions). In order to verify the dynamic capability of the instrument in real biological specimens, fluorescence changes of Fura-2 that were due to an intracellular flux of Ca2+ ions, and to an increase of [Ca2+]i (the intracellular Ca2+ concentration) have been recorded in Fura-2-loaded cultured cells of the line TE 671.

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Language(s): eng - English
 Dates: 1994-09-191994-08-031994-09-191995-03-01
 Publication Status: Published in print
 Pages: 10
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 Table of Contents: -
 Rev. Type: Peer
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Title: Pflügers Archiv: European Journal of Physiology
  Other : Pflügers Arch. Europ. J. Physiol.
Source Genre: Journal
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Publ. Info: Heidelberg : Springer-Verlag
Pages: - Volume / Issue: 429 (5) Sequence Number: - Start / End Page: 672 - 681 Identifier: ISSN: 0031-6768
CoNE: https://pure.mpg.de/cone/journals/resource/954925432380