Figure 4.

Analytical sensitivity of DNA sensors. (a) smFRET histograms of sensor-III for different concentrations of the target (0, 0.1, 0.2, 0.5, 1, 5, and 25 nM). The 0 nM data represent control experiment in the absence of the target. (b) Calibration curve of sensor-III depicting fraction of high-E_FRET population against target concentration. High-E_FRET population was determined from the two-peak Gaussian fitting of the histograms in Figure 3a. Inset displays the linear region of the full titration curve. R² value obtained from linear fitting was 0.95, and the limit of detection (LOD) was 36 pM, as determined using the equation, LOD = (3 × SD_blank)/slope, where SD_blank represents the standard deviation of high-E_FRET fraction in the absence of the target. (c) smFRET histograms of sensor-IV for different concentrations of the target. (d) Calibration curve of sensor-IV. Inset depicts the linear fitting region. R² value from linear fit was 0.97, and the detection limit was calculated to be 55 pM. Error bars in (b) and (d) represent the standard deviations (SD), n = 3. Each E_FRET histogram was prepared from 90–110 molecules.