Diagnostic importance of the two-stage factor VIII:C assay demonstrated by a case of mild haemophilia associated with His[sup 1954] → Leu substitution in the factor VIII A3 domain.

In some families with mild haemophilia higher results are obtained for factor VIII activity (FVIII:C) determined by one-stage assay than by two-stage or chromogenic assays. Amino-acid substitutions in the A1, A2 and A3 domains of factor VIII have been described in affected individuals with this phenotype. We describe a case of mild haemophilia A in which FVIII:C measured by one-stage assay was normal at 106%. However, FVIII:C levels measured by two-stage and chromogenic assays were 18% and 35% respectively. DNA analysis revealed a novel mutation in the A3 domain of factor VIII, His¹⁹⁵⁴ → Leu. In a molecular model of the FVIII A domains, His¹⁹⁵⁴ is placed in close proximity to two other mutations that have previously been shown also to be associated with one-stage/two-stage discrepancies. In this patient the diagnosis of haemophilia A would be missed if only the one-stage assay was used. [ABSTRACT FROM AUTHOR]