Prospect and Competence of Quantitative Methods via Real-time PCR in a Comparative Manner: An Experimental Review of Current Methods

There are numerous approaches dealing with relative and absolute quantitation. The methods differ in their efficiency assumption and applicability.

Current methodologies and rations used in qPCR quantification were compared in an experimental study of transgenic copy number determination of a monoclonal antibody Daclizumab.

With an inter and intra-methodical view, variations in relative and absolute quantification strategies were discretely extracted and compared to one another.

In relative quantification, six methods were studied and the ratios were computed relative to Glucagon as internal control. For Absolute quantification, the calculations were based on standard curve. Relative quantification considers the relative changes in expression levels while Absolute quantification relates the PCR signal to input copy number with a calibration curve.

The observed unevenness of the ratios in Relative approach pointed mainly to the efficiency changes and its calculation formula. Whereas results in Absolute approach strategies showed homogeneity which indicates the consistency of the calculation method.