Patent ID: 8744777

Claim:
A method for the quantification of a target nucleic acid in a test sample relative to a reference nucleic acid comprising: a) determining amplification efficiencies of the target nucleic acid and of the reference nucleic acid under defined amplification conditions, b) amplifying the target nucleic acid contained in the test sample and the reference nucleic acid contained in the test sample under the same defined reaction conditions, c) measuring the amplification of the target nucleic acid and of the reference nucleic acid in real time to determine amplified amounts of target nucleic acid and reference nucleic acid and calculating the ratio of the target nucleic acid and the reference nucleic acid in the test sample as a function of the amplification efficiencies determined in step a) and cycle number, d) amplifying the target nucleic acid contained in a calibrator sample and the reference nucleic acid contained in the calibrator sample under the same defined reaction conditions, e) measuring the amplification of the target nucleic acid from the calibrator sample and of the reference nucleic acid from the calibrator sample in real time to determine amplified amounts of the target nucleic acid and the reference nucleic acid from the calibrator sample and calculating the ratio of the target nucleic acid and the reference nucleic acid in the calibrator sample as a function of the amplification efficiencies determined in step a) and cycle number, and f) quantifying the target nucleic acid in the test sample relative to the reference nucleic acid as a function of a quotient of (i) said ratio of the target nucleic acid and the reference nucleic acid in the test sample and (ii) said ratio of the target nucleic acid and the reference nucleic acid in the calibrator sample.