PB1306 - Evaluation of a LAMP assay for detection of Prothrombin Allele and Factor V Leiden

Background: Prothrombin allele G20210A (PTA) and Factor V Leiden (FVL) are both inherited risk factors for venous thromboembolism and constitute part of a standard thrombophilia screen. Our laboratory evaluated the performance of reagents for PTA genotyping and FVL genotyping (La Car MDx, Belgium), tested using a Magnetic Induction Cycler (Bio Molecular System, Australia). These methods employ Loop Mediated Isothermal Amplification (LAMP). The main advantage of these LAMP methods is the ability to perform analysis on whole blood bypassing the need for DNA extraction typical required in PCR.

Aims: To compare the performance of LAMP methods against the current routine methods for genotyping PTA and FVL.

Methods: A mixture of citrate and EDTA samples stored at either 4 or -20 degrees and previously tested for PTA and FVL were included in this study. The predicate method was PTA and FVL reagents (Roche, UK) analysed on the LightCycler 2.0 (Roche, UK).

Results: Previously tested PTA whole blood samples comprising 117 wild-type, 61 heterozygous and 9 homozygous samples were evaluated. LAMP PTA genotyping demonstrated complete agreement with our current method. Previously tested FVL whole blood samples comprising 95 wild-type, 96 heterozygous and 21 homozygous samples were evaluated. LAMP FVL genotyping demonstrated complete agreement with our current method.

Conclusion(s): The LAMP method for PTA and FVL provides a more straightforward testing process by eliminating the need for time-consuming DNA extraction, thus shortening the preparation and with a reduced amplification/analysis time compared to LightCycler method. Overall, we had 100% agreement between PCR and LAMP methods for FVL and PTA genotyping.