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Updated: Jun 19, 2026
Development of a Quantitative Recombinase Polymerase Amplification Assay with an Internal Positive Control
Published on: March 30, 2015
Limitations of a calibrated, quantitative APC-R assay under routine conditions
Petra Jilma-Stohlawetz1, Katharina Lysy1, Raute Sunder-Plassmann1
1Department of Laboratory Medicine, Medical University of Vienna, Vienna, Austria.
Introduction:
The aim of this study was to evaluate the Hemoclot Quanti. V-L assay in various clinical conditions.
Methods:
We compared the Hemoclot Quanti.V-L assay with DNA testing and with the Pefakit assay in 60 normal (no mutation) vs carriers of the factor V (FV) Leiden mutation (56 heterozygous and three homozygous). We further investigated the interference of lupus anticoagulant on test results in normal and heterozygous individuals and of direct oral anticoagulants (DOACs) at trough and peak levels. Additionally, DOAC-Remove was tested in samples containing DOACs at peak levels. We further evaluated the influence of FV deficiency on this quantitative assay.
Results:
There was a 100% agreement between the Quant. V-L assay and DNA testing in 60 normal individuals. However, 1.85% of heterozygous and 33% of homozygous samples were falsely classified with the quantitative assay, and no misclassification was observed with the Pefakit assay. Lupus anticoagulant did not influence the test results of the quantitative assay. DOACs also interfered with test results in heterozygous patients, but this effect was prevented with the DOAC-Remove procedure. Even mild FV deficiency affected the test results of the quantitative assay in heterozygous patients leading either to misclassification or the need for subsequent PCR testing.
Conclusion:
The quantitative FV-L assay has several limitations, especially FV deficiency and the presence of DOACs have to be ruled out before running this quantitative assay.
