Factor V Null Mutation Affecting the Roche LightCycler Factor V Leiden Assay

Abstract

Although the role of factor V as a coagulation factor is more familiar, it has an equally important alternative role as a cofactor for protein C. Activated protein C (APC) is important in a naturally occurring anticoagulant pathway in which it cleaves factor V, thereby controlling the concentrations of factor V. The factor V Leiden mutation (1), which has a frequency of ∼1% in Caucasian populations and accounts for most cases of (APC) resistance, makes factor V resistant to cleavage by APC. Heterozygosity for the factor V Leiden mutation confers an increased lifelong relative risk for venous thrombosis, whereas homozygosity for the factor V Leiden mutation confers an even greater increased lifelong risk. Because of its high prevalence and association with thrombophilic disorders, a variety of assays have been developed to detect the G→A mutation at nucleotide 1691, codon 506, of the factor V gene, including assays based on use of the LightCycler™ (2)(3). In this report we present a case of an anomalous result obtained with the Roche LightCycler assay for factor V Leiden and discuss its implications. Recently, a 52-year-old female was diagnosed with deep venous thrombosis at our institution. As part of her assessment, she underwent a routine work-up for hypercoagulation. Her partial thromboplastin time and prothrombin time were normal before she was treated with anticoagulants. Measurements of protein C, protein S, and anti-thrombin III were deferred until she was finished with coumadin. Meanwhile, results from molecular diagnostics testing included a negative result for the G→A mutation at nucleotide 20210 in the prothrombin gene. However, an assay for the factor V Leiden mutation performed on the LightCycler showed an abnormal melting …