Abstract: A method for determining a threshold cycle number or time value in a nucleic acid amplification reaction comprising the steps of amplifying a nucleic acid sequence in a reaction mixture; at a plurality of different times during the amplification reaction, measuring at least one signal whose intensity is related to the quantity of the nucleic acid sequence in the reaction mixture; and storing signal values defining a growth curve for the nucleic acid sequence. A derivative of the growth curve is determined and the threshold cycle number or time value is calculated as a cycle number or time value corresponding to a characteristic (e.g., maximum, minimum, or zero-crossing) of the derivative.