What is the difference between Absolute Quantification and Relative Quantification in qPCR?
Posted March 25, 2022
Absolute quantification and relative quantification are two ways that are used to calculate the results of a qPCR experiment. There are a few significant differences between the two.
|Basis of differentiation||Absolute Quantification||Relative Quantification|
|What it determines||Expression levels in absolute numbers of copies||Fold changes in expression between two samples|
|Known/Unknown template amounts||The precise amount of the message or template used for the curve is known. The absolute quantification standard curve provides the final answer.||Although the template is known to contain the message of interest in high abundance, its absolute amount may or may not be known.|
|What the process involves||Unknowns are quantified based on a known quantity. A standard curve is first created. Then the unknowns are compared to the standard curve and a value is extrapolated.||Changes in gene expression in a given sample are analyzed relative to another reference sample such as an untreated control sample.|
|How the answer is determined||The standard curve provides the final answer.||The calibration curve result for the target gene is normalized to that of a housekeeping gene in the same sample. The normalized numbers are then compared between samples to obtain a fold change.|
Relative and Absolute Quantitative Real-Time PCR-Based Quantifications of hcnC and phlD Gene Transcripts in Natural Soil Spiked with Pseudomonas sp. Strain LBUM300