Single-Molecule-Sensitive FRET in Freely-Diffusing Attoliter Droplets

2015 
Fluorescence resonance energy transfer (FRET) from RNA confined in freely-diffusing attoliter aqueous droplets shows dramatic differences from that of RNA in solution. First, the use of droplet confinement is shown to substantially increase the signal-to-noise ratio of single-molecule sensitive measurements. However, a distinct shift in FRET is also observed. These differences can be attributed to a modified pH in the confining environment, which is a result of the well-documented autolysis of water and accumulation of hydroxide ions near the water interface. This outcome has implications for the use of droplets for protein crystallization, nanoparticle synthesis, biomicrofluidics, and analytical chemistry, where careful attention to the use of appropriate buffers or surfactants to control pH in the confined phase will be required. This work was funded by NSF MCB-0920139 and NSF DBI-1152386.
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