How to transfer a quantitative molecular diagnostic test to multiple qPCR platforms

Publication date: Available online 3 April 2018 Source:The Journal of Molecular Diagnostics Author(s): Claudia Gürtler, Mark Laible, Wolfgang Schwabe, Heike Steinhäuser, Xingmin Li, Shujin Liu, Kornelia Schlombs, Ugur Sahin Quantitative gene expression assays are increasingly used for diagnosis and research, but are often restricted to specific instrumentation. We propose a robust technical and statistical framework that enables transferring of established RT-qPCR assays across qPCR platforms without compromising analytical and clinical validity. The feasibility of our approach on MammaTyper®, an in vitro diagnostic assay which quantifies breast cancer biomarkers and dichotomizes results according to cut-off points, was tested. CFX96, ABI 7500 Fast, and Mx3000P were chosen as the candidate platforms whereas the LightCycler 480 II was used as a reference. Two instruments were used per platform, and tested initially for equivalence via Bland-Altman and Deming regression analyses. A method comparison approach was adapted to adjust cut-offs for the new systems, and evaluated the cross-platform agreement. Finally precision was estimated for each platform. The performance on the candidate devices was highly comparable to the reference platform with a 7 log quantification range and amplification efficiencies of 97% to 103%. The equivalence tests successfully pre-qualified instruments, preventing constant and proportional errors, and enabling reliable adjustments of cut-...
Source: The Journal of Molecular Diagnostics - Category: Pathology Source Type: research