Loop-mediated isothermal amplification (LAMP)-based method for detecting factor V Leiden and factor II G20210A common...

Area of Science:

• Molecular Diagnostics
• Genetics
• Thrombophilia Screening

Background:

• Rapid detection of Factor V Leiden and Factor II G20210A variants is crucial for thrombophilia testing due to high patient volumes.
• Current diagnostic methods often require DNA extraction, adding time and complexity.

Purpose of the Study:

• To validate a novel, CE-marked in vitro diagnostic (IVD) method for simultaneous detection of Factor V Leiden and Factor II G20210A variants.
• To assess the performance of a Loop-mediated isothermal AMPlification (LAMP) based duplex assay compared to a standard TaqMan SNP genotyping method.

Main Methods:

• A novel duplex Loop-mediated isothermal AMPlification (LAMP) assay was developed for simultaneous detection of Factor V Leiden and Factor II G20210A variants directly from whole blood samples.
• The novel LAMP method was validated against the routine TaqMan SNP genotyping method targeting genomic DNA.
• One hundred and eight patients undergoing thrombophilia testing were analyzed using both methods.

Main Results:

• The Loop-mediated isothermal AMPlification (LAMP) method demonstrated 100% concordance with the standard TaqMan SNP genotyping method.
• The assay correctly identified all genotypes for both Factor V Leiden and Factor II G20210A variants, including wild-type, heterozygotes, and homozygotes.
• The LAMP assay detected two double heterozygotes carrying both variants.

Conclusions:

• The novel Loop-mediated isothermal AMPlification (LAMP) assay provides accurate and reliable simultaneous detection of common thrombophilia variants.
• This validated IVD method offers comparable analytical performance to standard methods without the need for prior DNA extraction, enabling faster results.
• The duplex LAMP assay is a promising tool for efficient and rapid thrombophilia screening in clinical settings.