Illustration of cycle threshold concept for quantitative PCR (qPCR)

With successive cycles of PCR (cycle number on the X axis), the fluorescence signal from fluorescently tagged nucleotides incorporated into the PCR products increases (gray line). The cycle threshold (C_t, dotted black line) is set at 10^–1, where the signal is 10-fold greater than the background noise level of fluorescence seen with a negative control sample (blue line). In this example, the C_t of the test sample is 20 (dashed red arrow), meaning that 20 PCR cycles are required to reach the threshold fluorescence. A test sample with a greater amount of the starting material would reach the fluorescence threshold in fewer cycles and would have a lower C_t number.