High-Throughput Platform for Optoacoustic Probing of Genetically Encoded Calcium Ion Indicators

Functional optoacoustic (OA) imaging assisted with genetically encoded calcium ion indicators (GECIs) holds promise for imaging large-scale neuronal activity at depths and spatiotemporal resolutions not attainable with existing optical microscopic techniques. However, currently available GECIs optimized for fluorescence (FL) imaging lack sufficient contrast for OA imaging and respond at wavelengths having limited penetration into the mammalian brain. Here we present an imaging platform capable of rapid assessment and cross-validation between OA and FL responses of sensor proteins expressed in Escherichia coli colonies. The screening system features optimized pulsed light excitation combined with ultrasensitive ultrasound detection to mitigate photobleaching while further allowing the dynamic characterization of calcium ion responses with millisecond precision. Targeted probing of up to six individual colonies per second in both calcium-loaded and calcium-unloaded states was possible with the system. The new platform greatly facilitates optimization of absorption-based labels, thus setting the stage for directed evolution of OA GECIs. [Display omitted] •New platform for rapid screening of optoacoustic calcium ion indicators•Coregistered fluorescence and optoacoustic recordings from bacterial colonies•Sensitive imaging and selective probing in calcium-loaded and calcium-unloaded states•Assessment of protein photobleaching resistance to pulsed laser irradiation Analytical Chemistry; Bioengineering; Biomaterials