Improving positively tuned voltage indicators for faster kinetics and higher contrast

bioRxiv : the preprint server for biology
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Sungmoo Lee, Guofeng Zhang, Yukun Alex Hao, Richard H Roth, Yue Sun, Can Dong, Jun Zhu, Laura C Gomez, Ryan G Natan, Dongyun Jiang, Lan Xiang Liu, Guilherme Testa-Silva, Atsuki Hiramoto, Botond Roska, Daniel Feldman, Na Ji, Thomas R Clandinin, Lisa Giocomo, Jun Ding, Michael Z Lin

bioRxiv [Preprint]. 2026 May 4:2024.06.21.599617. doi: 10.1101/2024.06.21.599617.

ABSTRACT

Existing positively tuned ASAP-family voltage indicators such as ASAP4e exhibit superior photostability compared to the negatively tuned ASAP3 and ASAP5, but signal-to-noise ratios for spike detection remained similar to ASAP3. To improve spike detection by positively tuned ASAP indicators, we performed multiple rounds of structure-guided saturation mutagenesis of an ASAP4e predecessor while screening directly for faster responses. One variant, ASAP6c, demonstrated ~100% increases in fluorescence in response to single action potentials in both one-photon and two-photon imaging in vivo, achieving a 3-fold improvement in signal-to-noise ratios over ASAP4e.

PMID:42146340 | PMC:PMC13174376 | DOI:10.1101/2024.06.21.599617