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ReaLeyden Test

Set of reagents for determining factor V resistance to protein C (ReaLeyden test) according to TU 9398-034-05595541-2011 is intended for determining the resistance of activated factor V (factor V Leiden) to the action of activated Protein C in human blood plasma using the clotting method on all types of coagulometers.

The anticoagulant system of protein C (protein C, protein S, thrombomodulin, thrombin, protein C inhibitor) plays a crucial role in regulating the coagulation activity of blood plasma. The system primarily acts to inhibit factors VIIIa and Va and to inactivate plasminogen activator inhibitor (PAI-1). Abnormalities in the system can lead to thromboses of various locations. A new form of inherited thrombophilia has been characterized, based on the presence of the so-called V factor Leiden in patients. The essence of this form is that V factor Leiden (mutant factor V of the blood coagulation system) is resistant to the action of activated protein C, leading to increased blood coagulation through internal and external mechanisms and increasing the frequency of thromboses, including peripheral vein thrombosis, myocardial infarctions, pulmonary embolisms, retinal vein thrombosis, and others.

Method Principle: Incubation of normal plasma with an activator (venom of Agkistrodon contortrix) activates endogenous protein C and protein S, leading to prolonged clotting time of normal plasma in the activated partial thromboplastin time (APTT) test. Without the addition of the APTT activator, clotting time in the same plasma remains unchanged. In the plasma of patients with a deficiency in the protein C system or the presence of mutant factor V (Leiden), the prolongation of APTT upon the addition of the activator is less pronounced than in the norm.

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Set composition:

  • APTT (Activated Partial Thromboplastin Time) reagent, lyophilized (2 ml/vial) – 1 pc.;
  • APTT reagent with protein C activator, lyophilized (2 ml/vial) – 1 pc.;
  • Plasma calibrator, lyophilized (1 ml/vial) – 1 pc.;
  • Factor V-deficient substrate plasma (1 ml/vial) – 4 pcs.;
  • 0.025 M calcium chloride solution (5 ml/vial) – 1 pc.

The kit is designed for conducting 20 determinations on all types of coagulometers.

Results interpretation: The test results are conventionally expressed as the Normalized Ratio (NR). In healthy individuals, NR is 1.1 ± 0.3. An NR value below 0.8 suggests the possible presence of mutant factor V (Leiden), resistant to the action of activated protein C. False-positive results may occur when analyzing plasma samples from patients:

  • with elevated heparin levels;
  • with systemic lupus erythematosus;
  • when using direct-acting anticoagulants (INR above 3.0). Final confirmation of the presence of the Leiden mutation can be made through PCR diagnostics.

Normal and pathological values of the Normalized Ratio should be monitored using control plasmas from NPO RENAM: Plasma control with normal and reduced levels of protein C (Plasma-Protein C) (code PFA-6/7).