Figure 4.
Microscope images of negative (a,b) and positive (c,d) droplet samples encapsulating an engineered E. coli strain containing the synthetic blaCTX-M-9 target gene before (a,c) and after (b,d) thermocycling. Cy5 fluorescence images were recorded to measure droplet SNR for different replicates. (Same intensity range for all panels, scaled to visualize trace fluorescent signal in negative control droplets. Scalebar = 500 μm). e) Quantitative intensity measurements for droplets in panels B (negative control) and D (positive control), where overall SNR averages 2.87 (defined as 90th % positive droplet intensity divided by average negative droplet intensity), (n = 473 individual droplets). The optimal conditions used in this experiment were 10% whole blood, 40 PCR cycles, 2:1 primer-to-probe ratio (1μM primer, 500 nM probe), 5mg/ml BSA, 0.2% NP-40, and a 70 μl mineral oil overlay over droplets during thermocycling.