Fitting the whole curve and determining the exponential phase. The fluorescent data from a typical sample in this study (filled circles) are plotted. (**A**) Four four-parameter S-shaped models, Logistic (red), Sigmoid (blue), Gompertz (brown), and Chapman (purple) model together with an ideal three-parameter simple exponent model (dashed line) were fitted to the PCR kinetic curve. The symbol *a* is the difference between the maximum fluorescence and the ground fluorescence; *y*_{0} is the ground fluorescence. **Inset:** Magnified view of the exponential phase between SPE and EPE. (**B**) Determining the start point. The plot of sample in (A) is expanded to allow visualization of the ground phase. When the baseline is calculated from the entire ground phase including two points (blue) with higher noise, a later outlier is identified as the start point of exponential phase (blue arrow). The efficiency calculated by this method is an overestimation (*E* = 1.397, blue) when compared with the efficiency estimated by standard curve (*E*_{std} = 0.9747). After deletion of the high noise cycles based on subjective judgment, a refined baseline (red) identifies an earlier outlier and generates an improved efficiency estimate (*E* = 0.936, red). Our noise level based SPE algorithm defined the start point without making assumptions about the baseline, which resulted in a closer efficiency (*E* = 0.942, black arrow) to *E*_{std} even from this single reaction. Using this method, the final averaged efficiency for this gene (*E*_{Miner} = 0.9630) is very close to the efficiency estimated by the standard curve method. (**C**) UBS and OBS: UBS (red) will result when cycles with high noise in the ground phase exist on the upper side of the ideal baseline (dashed line), resulting in subtracting smaller values for later cycles after the ground phase. In the opposite, OBS (blue) due to the existence of the points with high noise on the lower side of the ideal baseline results in subtracting too much for later cycles after the ground phase. The sample is the same as used in (B).

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