A new clotting method for quantitating Factor V-Leiden in plasma


Hyphen Biomed presents a quantitative clotting method for Factor V-Leiden (FV L)  measurement in citrated plasma, based on FV-L’s resistance to  Activated Protein C (APC). Normal FV is not measured with the assay, as it is totally inhibited by APC, whilst FV-L keeps its clotting activity.



Diluted plasma is mixed with a purified clotting factor mixture, in a constant and optimised concentration, (R1 : Fibrinogen, Prothrombin, Protein S and APC). Purified FXa, with phospholipids (R2), is then added. Coagulation is initiated by the addition of calcium (Ca2+) and the clotting time (CT) is measured. The CT obtained is inversely proportional to the FV-L concentration. An inverse linear relationship is obtained between the CT and the FV-L concentration expressed as a percentage (%).Calibration

A standard curve is constructed using a FV-L heterozygous plasma pool  as a calibrator.


  • A totally quantitative assay for the measurement of FV-L concentrations on citrated plasma is presented.
  • Only one clotting test is required (= no problem of result interpretation).
  • Excellent discrimination (for both clotting times and FV-L concentrations) between normals, heterozygous and homozygous patients (for the R506Q mutation ).
  • Patients with low concentrations of total FV clotting activity (<25%) must be identified, and the diagnosis confirmed by comparing FV-L and total FV clotting activity (normals < 0.1).
  • No interference of Heparin or Dicoumarol therapy.
  • Easy to perform, cost effective and reliable assay, fully automatable on laboratory instruments.


Factor V-Leiden concentrations in normals and patients carrying the FV-L (R506Q) mutation:

Patients FVL Conc. (%) CT* (sec)
NORMAL(N=30)  <10  >100
HETEROZYGOUSZ(N=8) 35 – 60 50 – 70
HOMOZYGOUS (N=2) >100  <40CT

*CT may vary from lot to lot, but FV-L concentrations are accurately determined by using the calibration curve specific for each lot and each run.


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