Browsing by Author "Ballanyi, Klaus"

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A long Stokes shift red fluorescent Ca2+ indicator protein for two-photon and ratiometric imaging
(Nature Publishing Group, 2014-10) Wu, Jiahui; Abdelfattah, Ahmed S.; Miraucourt, Loïs S.; Kutsarova, Elena; Ruangkittisakul, Araya; Zhou, Hang; Ballanyi, Klaus; Wicks, Geoffrey; Drobizhev, Mikhail; Rebane, Aleksander; Ruthazer, Edward S.; Campbell, Robert E.
The introduction of calcium ion (Ca2+) indicators based on red fluorescent proteins (RFPs) has created new opportunities for multicolour visualization of intracellular Ca2+ dynamics. However, one drawback of these indicators is that they have optimal two-photon excitation outside the near-infrared window (650–1,000 nm) where tissue is most transparent to light. To address this shortcoming, we developed a long Stokes shift RFP-based Ca2+ indicator, REX-GECO1, with optimal two-photon excitation at <1,000 nm. REX-GECO1 fluoresces at 585 nm when excited at 480 nm or 910 nm by a one- or two-photon process, respectively. We demonstrate that REX-GECO1 can be used as either a ratiometric or intensiometric Ca2+ indicator in organotypic hippocampal slice cultures (one- and two-photon) and the visual system of albino tadpoles (two-photon). Furthermore, we demonstrate single excitation wavelength two-colour Ca2+ and glutamate imaging in organotypic cultures.
A sensitive and specific genetically-encoded potassium ion biosensor for in vivo applications across the tree of life
(Public Library of Science, 2022-09) Wu, Sheng-Yi; Wen, Yurong; Serre, Nelson B. C.; Heiede Laursen, Cathrine Charlotte; Grostøl Dietz, Andrea; Taylor, Brian R.; Drobizhev, Mikhail; Molina, Rosana S.; Aggarwal, Abhi; Rancic, Vladimir; Becker, Michael; Ballanyi, Klaus; Podgorski, Kaspar; Hirase, Hajime; Nedergaard, Maiken; Fendrych, Matyáš; Lemieux, M. Joanne; Eberl, Daniel F.; Kay, Alan R.; Campbell, Robert E.; Shen, Yi
Potassium ion (K+) plays a critical role as an essential electrolyte in all biological systems. Genetically-encoded fluorescent K+ biosensors are promising tools to further improve our understanding of K+-dependent processes under normal and pathological conditions. Here, we report the crystal structure of a previously reported genetically-encoded fluorescent K+ biosensor, GINKO1, in the K+-bound state. Using structure-guided optimization and directed evolution, we have engineered an improved K+ biosensor, designated GINKO2, with higher sensitivity and specificity. We have demonstrated the utility of GINKO2 for in vivo detection and imaging of K+ dynamics in multiple model organisms, including bacteria, plants, and mice.