Two-Stage Clotting Factor VIII:C

Coagulation

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Description

The two-stage clotting FVIII was first introduced in the 1950s. Activated FVIII (FVIIIa), along with FIXa, is a cofactor which activates FX to FXa in the tenase complex of coagulation. The concentration of FVIII(a) is made the rate limiting step in the assay and FXa generated is proportional to this. The FXa generated is measured by time to clot.

Test plasma is incubated with aluminium hydroxide (alumina) to remove vitamin K-dependent clotting factors (FII, FVII, FX, FIX) and FVIIIa. This is then mixed with activated serum (provides FIX, FX, FXIa), calcium, phospholipids and FV. FXa is generated. Normal plasma is added (provides prothrombin and fibrinogen) to allow clot formation.

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Indications

Factor VIII (FVIII) is an essential coagulation protein. A reduction or absence of FVIII results causes the X-linked bleeding disorder, haemophilia A. A reduction in FVIII may also be observed in patients with von Willebrand disease (VWD). FVIII activity (FVIII:C) may be measured by one-stage clotting, two-stage clotting or chromogenic assays.

Approximately one third of patients with the mild form of haemophilia A will demonstrate significantly different FVIII:C when measured by one-stage FVIII compared to the two-stage or chromogenic FVIII. This is known as assay discrepancy. Half of patients with assay discrepancy will have a two-fold or lower one-stage FVIII than two-stage or chromogenic FVIII and approximately half will have a two-fold or lower two-stage or chromogenic FVIII than one-stage FVIII. There is a genetic link to both versions of FVIII assay discrepancy.

The two-stage assay is only performed in the diagnosis of mild haemophilia A to determine the presence of FVIII assay discrepancy.

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Sample Type

Plasma (Citrate Blue) x 2.

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Reference Range

Adult reference range is indicated on the report.

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Turnaround Time

Within 14 days (or on demand)

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Testing Frequency

As required.

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References

Bowyer AE, Van Veen JJ, et al. Specific and global coagulation assays in the diagnosis of discrepant mild hemophilia A. Haematologica. 2013; 98: 1980-7
Bowyer AE, Goodeve A, et al. p.Tyr365Cys change in factor VIII: haemophilia A, but not as we know it. Br J Haematol. 2011; 154: 618-25
Peyvandi F, Oldenburg J, et al. A critical appraisal of one-stage and chromogenic assays of factor VIII activity. J Thromb Haemost. 2016; 14: 248-61

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See Also

Chromogenic Factor VIII

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