The precise control of biochemical reactions in live cells is a long-sought goal for researchers. Conventional approaches such as chemical treatment and genetic methods lack spatial and temporal precision. Current laser-based approaches such as optical tweezers, laser ablation, and optogenetics have slow temporal responses, i.e. requiring pre-imaging and manual target selection and treatment. For live biological samples, highly dynamic processes together with highly mobile organelles make pre-imaging and selection difficult for meaningful capturing and control of complex biochemical consequences. To address this, we developed a real-time precision opto-control (RPOC) technology that during imaging can control dynamic cellular processes simultaneously and automatically for real-time analysis. By comparing generated optical signals during laser scanning with tunable preset voltage values through a comparator circuit, we can control the 1<sup>st</sup> order output of a fast optical shutter with a nanosecond response time to only activate the manipulation laser at the pixels with the species of interest with a feedback loop faster than the pixel dwell time during imaging. RPOC allows us to control chemical processes with high spatial and temporal accuracy and high chemical selectivity. By coupling multiple comparator circuits in tandem, we can achieve higher-order voltage selection processes such as bandpass filters and digital logic for higher chemical selectivity. To demonstrate the capability of RPOC in live cells, we successfully controlled the states of photochromic molecules at different parts of the cell. Paired with photoswitchable inhibitors, we also controlled intercellular organelle dynamics via inhibiting polymerization of microtubule.

Imaging and manipulating chemicals in live cells using real-time precision opto-control

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