Positional analysis via C
. (A) Profile position is defined by the fractional cycle (designated by the red arrows) at which reaction fluorescence reaches half of maximal (Fmax). Calculated using the inlaid equation, C1/2 is derived from sigmoidal analysis of each individual amplification profile, rather than from defining a common fluorescence threshold as is done for determining Ct. C1/2 thus provides positional information that is unique to each individual amplification profile. (B) Identical to the standard curve analyses presented in Figures 1B and 2B, amplification efficiency is derived from the slope and the number of amplicon molecules at C1/2 (NC1/2) derived from the intercept.