Diagnostic importance of the two-stage factor VIII:C assay demonstrated by a case of mild haemophilia associated with His1954 → 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, His(1954)-->Leu. In a molecular model of the FVIII A domains, His(1954) 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.