A genetically encoded far-red fluorescent calcium ion biosensor derived from a biliverdin-binding protein
- Hashizume, Rina
- Fujii, Hajime
- Mehta, Sohum
- Ota, Keisuke
- Qian, Yong
- Zhu, Wenchao
- Drobizhev, Mikhail [ Montana State University: Microbiology & Cell Biology ]
- Nasu, Yusuke
- Zhang, Jin
- Bito, Haruhiko
- Campbell, Robert E.
Far-red and near-infrared (NIR) genetically encoded calcium ion (Ca2+) indicators (GECIs) are powerful tools for in vivo and multiplexed imaging of neural activity and cell signaling. Inspired by a previous report to engineer a far-red fluorescent protein (FP) from a biliverdin (BV)-binding NIR FP, we have developed a far-red fluorescent GECI, designated iBB-GECO1, from a previously reported NIR GECI. iBB-GECO1 exhibits a relatively high molecular brightness, an inverse response to Ca2+ with ΔF/Fmin = −13, and a near-optimal dissociation constant (Kd) for Ca2+ of 105 nM. We demonstrate the utility of iBB-GECO1 for four-color multiplexed imaging in MIN6 cells and five-color imaging in HEK293T cells. Like other BV-binding GECIs, iBB-GECO1 did not give robust signals during in vivo imaging of neural activity in mice, but did provide promising results that will guide future engineering efforts.