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Proceedings Paper

Auto-luminescent genetically encoded ratiometric indicator for real-time Ca2+ imaging at the single cell level
Author(s): Kenta Saito; Kentaro Kobayashi; Takeharu Nagai
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Paper Abstract

Efficient bioluminescence resonance energy transfer (BRET) from a bioluminescent protein to a fluorescent protein with high fluorescent quantum yield has been utilized to enhance luminescence intensity, allowing single-cell imaging in near real time without external light illumination. We have applied this strategy to develop an autoluminescent Ca2+ indicator, BRAC, which is composed of Ca2+-binding protein, calmodulin, and its target peptide, M13, sandwiched between a yellow fluorescent protein variant, Venus, and an enhanced Renilla luciferase, RLuc8. With this BRAC, we succeeded visualization of Ca2+ dynamics at the single-cell level with temporal resolution at 1 Hz. Moreover, BRAC signals were acquired by ratiometric imaging capable of canceling out Ca2+-independent signal drifts due to change in cell shape, focus shift, etc. Taking advantage of the bioluminescence imaging property that does not require external excitation light, BRAC might become a powerful tool applicable in conjunction with so-called optogenetic technology by which we can control cellular and protein function by light illumination.

Paper Details

Date Published: 23 December 2011
PDF: 6 pages
Proc. SPIE 8204, Smart Nano-Micro Materials and Devices, 820403 (23 December 2011); doi: 10.1117/12.905065
Show Author Affiliations
Kenta Saito, Hokkaido Univ. (Japan)
Kentaro Kobayashi, Hokkaido Univ. (Japan)
Takeharu Nagai, Hokkaido Univ. (Japan)
Japan Science and Technology Agency (Japan)

Published in SPIE Proceedings Vol. 8204:
Smart Nano-Micro Materials and Devices
Saulius Juodkazis; Min Gu, Editor(s)

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