What are the methods commonly used to quantify the results obtained by real-time RT-PCR?

The methods commonly used to quantify results obtained by real time RT-PCR include the standard curve method and the comparative threshold method. 

1. The standard curve method is a technique used to quantify the concentration of mRNA targets of interest. This curve serves as a reference for determining the concentrations of mRNA targets with unknown concentrations. Initially, a standard curve is established using RNA of known concentrations; using RNA standards with known concentrations enables the generation of absolute copy number data. Alternatively, other nucleic acid samples such as in vitro generated ssDNA, purified plasmid dsDNA, or any cDNA sample expressing the target gene can be used to construct the standard curve.
2. The comparative threshold method compares the Ct values of samples of interest with a control or calibrator, typically a non-treated sample or RNA from normal tissue. Both the reference (calibrator) and the samples being tested are adjusted or standardized using a housekeeping gene. [delta][delta]Ct is calculated by subtracting the Ct value of the calibrator from the Ct value of the sample, after both are normalized to the housekeeping gene. For this calculation to be valid, the amplification efficiencies of the target and the endogenous reference should be relatively equal. If a suitable housekeeping gene cannot be identified with similar amplification efficiency to the target gene, then the standard curve method should be used.