Figure 2.

Shrinkage-based droplet analysis of bacterial growth. (A) Fluorescence-enabled, droplet-based microfluidic screening pipeline. Bacteria are individualized into 20 pL of water-in-oil droplets in the presence of the Alexa Fluor 488 fluorophore at 5 μg/mL final concentration (left), collected, and incubated from 2 to 20 h at the chosen temperature (middle). Droplets are then reinjected into a fluorescence-activated droplet sorting device (right) in which the fluorescence of each droplet is analyzed and used for sorting droplets displaying the fluorescence profile of interest. (B) Typical fluorescence profile obtained after incubation. Since not all droplets are occupied by bacteria, at least two distinct populations of droplets can be detected after incubation. Empty droplets conserve their original volume (unshrunk droplets) and show moderate fluorescence (∼20 000 rfu). Droplets in which bacterial growth took place experience shrinkage (shrunk droplets), leading to an increase in fluorophore concentration and thus in fluorescence intensity (∼35 000 rfu). Computing the fluorescence ratio of both populations yields the shrinkage index (here, shrinkage index ∼ 1.75). The result can be confirmed by epifluorescence microscopy (inset).