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Proceedings Article

Development of ultra-sensitive Ca2+ indicators, yellow cameleon-nanos

[+] Author Affiliations
Takeharu Nagai, Kazuki Horikawa

Hokkaido Univ. (Japan) and Japan Science and Technology Agency (Japan)

Tomoki Matsuda, Kentaro Kobayashi

Hokkaido Univ. (Japan)

Proc. SPIE 7910, Reporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications III, 79100G (February 11, 2011); doi:10.1117/12.871202
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From Conference Volume 7910

  • Reporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications III
  • Samuel Achilefu; Ramesh Raghavachari
  • San Francisco, California, USA | January 22, 2011

abstract

There are so many Ca2+ indicators such as Oregon green 488 BAPTA-1 and cameleon, all of which have moderate Ca2+ affinities (K d > 150 nM). These indicators successfully report changing in Ca2+ concentration in most of cells. However, it is known that some cells are estimated to have a very low resting [Ca2+] and display very small ▵[Ca2+]s, which is below the detection limit of the indicators. To develop high-affinity indicators, we modified a FRET-based Ca2+ indicator, yellow cameleon (YC) 2.60 (K d = 100 nM) because of the brightness and the large dynamic range. By engineering the Ca2+ sensing domain, we obtained a series of ultra-sensitive Ca2+ indicators. Their high Ca2+ affinities (Kd = 15, 30, and 50 nM) enabled detection of subtle Ca2+ transients associated with spontaneous network activity in zebrafish. Our measurements revealed that both the resting Ca2+ level and the amplitude of Ca2+ transients significantly differed by cell type and stimulation, indicating that the selection of indicators with the appropriate K d is essential for successful in vivo Ca2+ imaging. A lineup of such indicators with finely tuned Kd values optimized to detect [Ca2+] from 10 nM to 100 nM would enable precise and reliable Ca2+ imaging.

© (2011) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE). Downloading of the abstract is permitted for personal use only.
Citation

Takeharu Nagai ; Kazuki Horikawa ; Tomoki Matsuda and Kentaro Kobayashi
"Development of ultra-sensitive Ca2+ indicators, yellow cameleon-nanos", Proc. SPIE 7910, Reporters, Markers, Dyes, Nanoparticles, and Molecular Probes for Biomedical Applications III, 79100G (February 11, 2011); doi:10.1117/12.871202; http://dx.doi.org/10.1117/12.871202


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