Method for the efficiency-corrected real-time quantification of nucleic acids

Method for the quantification of a target nucleic acid in a sample relative to a reference nucleic acid comprising the following steps:

• a) determination of the amplification efficiencies of the target nucleic acid and of the reference nucleic acid under defined amplification conditions
• b) amplification of the target nucleic acid contained in the sample and of the reference nucleic acid contained in the 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
• d) calculation of the original ratio of target nucleic acid and reference nucleic acid in the sample by correction of the ratio derived from step c) with the aid of the amplification efficiencies determined in step a).

Method for the quantification of a target nucleic acid in a sample relative to a reference nucleic acid comprising the following steps:

• a) determination of the amplification efficiencies of the target nucleic acid and of the reference nucleic acid under defined amplification conditions
• b) amplification of the target nucleic acid contained in the sample as well as of the reference nucleic acid contained in the sample under the same defined amplification conditions.
• c) measurement of the amplification of the target nucleic acid and of the reference nucleic acid in real time.
• d) determining a defined signal threshold value
• e) determining the cycle numbers at which the signal threshold value is exceeded in each case during the amplification of target nucleic acid and reference nucleic acid
• f) calculating the original ratio of target nucleic acid and of reference nucleic acid in the sample according to the following formula
• N(T)0/N(R)0=E(R)n(R)/E(T)n(T), in which
• N(T)0=the original amount of target DNA present in the sample
• N(R)0=the original amount of reference DNA present in the sample
• E(R)=the amplification efficiency of the reference nucleic acid
• n(R)=the cycle number of the reference nucleic acid measured in step e)
• E(T)=the amplification efficiency of the target nucleic acid
• n(T)=the cycle number of the target nucleic acid measured in step e).

Method for the quantification of a target nucleic acid in a sample comprising the following steps:

• a) determination of the amplification efficiencies of the target nucleic acid and of an internal or external standard under defined amplification conditions
• b) amplification of the target nucleic acid contained in the sample and of the internal or external standard under the same defined reaction conditions
• c) measuring the amplification of the target nucleic acid and of the standard in real time
• d) calculating the original copy number in the sample by correction of the copy number derived from step c) with the aid of the amplification efficiencies determined in step a).

Method for the quantification of a target nucleic acid in a sample comprising the following steps:

• a) determination of the amplification efficiencies of the target nucleic acid and of an internal or external standard under defined amplification conditions
• b) amplification of the target nucleic acid contained in the sample as well as of the internal or external standard under the same defined reaction conditions.
• c) measurement of the amplification of the target nucleic acid and of the standard in real time.
• d) determining a defined signal threshold value
• e) determining the cycle numbers at which the signal threshold value is exceeded during the amplification of target nucleic acid and standard
• f) determining the original copy number N(T)0 of target nucleic acid in the sample according to the formula
• N(T)0=N(S)0*E(S)n(s)/E(T)n(T), in which
• N(S)0=the original amount of standard used
• E(S)=the amplification efficiency of the standard
• n(S)=the cycle number of the standard measured in step e)
• E(T)=the amplification efficiency of the target nucleic acid
• n(T)=the cycle number of the target nucleic acid measured in step e).