fig1

Figure 1. Preparation and Serial Dilution of Exogenous Spike-in Control. (A) Serial dilution instructions for creating a 25 µM stock solution of cel-miR-39-3p (miR-39) spike-in. First, a 0.5 µM solution of miR-39 spike-in is prepared by performing a 1:50 dilution in water. The actual stock concentration (in ng/μL) is then measured by a Qubit Fluorometer and microRNA Concentration Kit. The concentration of the stock solution is adjusted to 1,000 ng/μL. Serial dilutions of the miR-39 spike-in are prepared in water as shown. All dilutions are prepared by mixing 1 part of miR-39 spike-in solution with 9 parts water, vortexing (30 s) and resting on ice (30 s) 5 times before subsequent dilution. (B) Representative 1-Demensional analysis of 01-0.0001 ng/μL and the respective No Template Control (NTC). The green dots represent VIC positive (miR-39) particles, while the grey dots represent droplets with no fluorescence. (C) Ratio of miR-39 positive to total events. Results represent triplicate measurements, ±standard error of the mean (SEM). (D) Measured concentrations of miR-39 (copy number/μL), where the solid line is the median, and the bar represents the interquartile range.