Abstract
Cellular heterogeneity has presented a significant challenge in the studies of biology. While most of our understanding is based on the analysis of ensemble average, individual cells may process information and respond to perturbations very differently. Presented here is a highly sensitive platform capable of measuring enzymatic activity at the single-cell level. The strategy innovatively combines a rolling circle-enhanced enzyme activity detection (REEAD) assay with droplet microfluidics. The single-molecule sensitivity of REEAD allows highly sensitive detection of enzymatic activities, i.e. at the single catalytic event level, whereas the microfluidics enables isolation of single cells. Further, confined reactions in picoliter-sized droplets significantly improve enzyme extraction from human cells or microorganisms and result in faster reaction kinetics. Taken together, the described protocol is expected to open up new possibilities in the single-cell research, particularly for the elucidation of heterogeneity in a population of cells.
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Tesauro, C., Frøhlich, R., Stougaard, M., Ho, YP., Knudsen, B.R. (2015). Microfluidics-Enabled Enzyme Activity Measurement in Single Cells. In: Singh, A., Chandrasekaran, A. (eds) Single Cell Protein Analysis. Methods in Molecular Biology, vol 1346. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2987-0_14
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DOI: https://doi.org/10.1007/978-1-4939-2987-0_14
Publisher Name: Humana Press, New York, NY
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