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bioRxiv · 10.64898/2026.01.20.700092

Bright calcium-modulated bioluminescent indicators for activity imaging and red photon-assisted synaptic transmission

Zhao, Y.·Porta-de-la-Riva, M.·Lee, S.·Wu, Y.·Klein, M.·Hall, M.·Su, Y.·Encell, L.·Kirkland, T.·Krieg, M.·Lin, M.
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Abstract

Bioluminescent calcium sensors have unique uses in neuroscience and neuroengineering, enabling noninvasive imaging of neuronal activity and contactless activation of opsin-expressing neurons. However, the speed, range, and robustness of non-invasive imaging and rewiring or neuronal activity are limited by the radiance and dynamic range of existing bioluminescent calcium sensors. Here, we report the stepwise engineering of improved cyan-excitable red fluorescent proteins (mCyRFP4 and dCyRFP4), an improved red bioluminescent protein based on NanoLuc and CyRFP4 (Antares3), and an improved bioluminescent calcium sensor based on Antares3 (CaMBI3). Antares3 is 3-fold brighter than its predecessor, while CaMBI3 responds with an overall dynamic range of 24-fold, enabling high-sensitivity detection of calcium dynamics in muscle and neuronal tissues in vivo. Finally, a CaMBI3 variant and a red-shifted opsin ChRmine enabled red photon-assisted synaptic transmission in C. elegans. CaMBI3 thus facilitates genetically targeted non-invasive imaging and rewiring of neural activity in living animals.

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Zhao, Y., Porta-de-la-Riva, M., Lee, S., Wu, Y., Klein, M., Hall, M., Su, Y., Encell, L., Kirkland, T., Krieg, M., Lin, M.. 2026-01-23. Bright calcium-modulated bioluminescent indicators for activity imaging and red photon-assisted synaptic transmission. https://doi.org/10.64898/2026.01.20.700092

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