Abstract
Cells typically respond to chemical or physical perturbations via complex signaling cascades which can simultaneously affect multiple physiological parameters, such as membrane voltage, calcium, pH, and redox potential. Protein-based fluorescent sensors can report many of these parameters, but spectral overlap prevents more than ~4 modalities from being recorded in parallel. Here we introduce the technique, MOSAIC, Multiplexed Optical Sensors in Arrayed Islands of Cells, where patterning of fluorescent sensor-encoding lentiviral vectors with a microarray printer enables parallel recording of multiple modalities. We demonstrate simultaneous recordings from 20 sensors in parallel in human embryonic kidney (HEK293) cells and in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), and we describe responses to metabolic and pharmacological perturbations. Together, these results show that MOSAIC can provide rich multi-modal data on complex physiological responses in multiple cell types.
Highlights
Cells typically respond to chemical or physical perturbations via complex signaling cascades which can simultaneously affect multiple physiological parameters, such as membrane voltage, calcium, pH, and redox potential
Each fluorescent sensor was validated in iPSC-derived cardiomyocytes and many were validated in human embryonic kidney 293T (HEK293) cells
For sensors targeted to subcellular locations, we confirmed trafficking with a live-cell stain for mitochondria or endoplasmic reticulum (Methods)
Summary
Cells typically respond to chemical or physical perturbations via complex signaling cascades which can simultaneously affect multiple physiological parameters, such as membrane voltage, calcium, pH, and redox potential. We demonstrate simultaneous recordings from 20 sensors in parallel in human embryonic kidney (HEK293) cells and in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), and we describe responses to metabolic and pharmacological perturbations. Together, these results show that MOSAIC can provide rich multi-modal data on complex physiological responses in multiple cell types. When a cell responds to external cues, multiple second messengers may be involved, and the cell state can change in many ways To follow these complex, correlated dynamics, it is desirable to record from many fluorescent sensors simultaneously. We demonstrate simultaneous recording of action potentials and calcium transients in the cytosol, mitochondria, and endoplasmic reticulum (ER) in a synchronized cardiomyocyte syncytium. (5) Cellular microarrays minimize the usage of cells and reagents, which can be critical for expensive primary or stem cell-derived samples
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