FIGURE 4.

Improved droplet separation with the addition of restriction enzyme in the MuSIC ddPCR assay. (A) The integration of BamHI restriction enzyme digestion in MuSIC ddPCR was assessed using a panel of EHEC strains with various serological profiles. Association of EHEC virulence targets was assessed by calculating a normalized value for linkage, based on the values provided by the QuantaSoft^TM software (Percent linkage). Only minor differences in the proportion of droplets with amplification of both stx/eae targets was observed in assays with restriction enzyme (red bars) compared to reactions without restriction enzyme (blue bars). Error bars represent the standard deviation of 4 replicates. (B) An increase in the mean fluorescence amplitude of positive droplets for the stx assay (left panel) relative to assays conducted without the enzyme was observed. The use of the restriction enzyme did not seem to affect amplification of the eae target (right panel).