Assays for von Willebrand Factor Functional Activity: A UK NEQAS Survey

In most laboratories, the diagnosis of von Willebrand’s disease hinges largely on the results of assays of FVIII:C, VWF:Ag and VWF:RiCof. The VWF:RiCof assay is a functional assay which, although not perfect, is thought to mirror the functional activity of von Willebrand factor in respect of its interaction with platelet membrane GP1b/IX. In a clinical context the importance of the VWF:RiCof assay is that it permits the identification of type 2A vWD which is characterized by disproportionately low levels of vWF:RiCof relative to vWF:Ag. This distinction is of some clinical importance since the bleeding manifestations of individuals with type 2A vWD are often more marked than those of individuals with the commoner type 1 variant. The haemostatic response to DDAVP is also less predictable in these patients and clotting factor concentrates can be required for their management. In 1997, Murdock et al. described a new ELISA assay which involved the use of a monoclonal antibody and which recognised an epitope on von Willebrand factor important in its interaction with GP1b. The authors considered that the ELISA provided an assay for vWF functional activity and they concluded that when used in conjunction with an ELISA for vWF:Ag and vWF multimer analysis, it provided a reliable method for the laboratory diagnosis of vWD. Information derived from returns from participants in the UK National External Quality Assessment Scheme (UK NEQAS) in Blood Coagulation indicates that an increasing number of UK laboratories have adopted the ELISA assay by Murdock et al (1997) for the diagnosis of vWD. In a recent survey, undertaken in September 1997, it was performed by 27 of the 108 laboratories registered for vWF activity assays. From data derived from UK NEQAS surveys it is now clear that there can be significant differences between vWF:RiCof results and those obtained by the ELISA method for vWF activity. For example, the median vWF “activity” obtained by ELISA on a lyophilised plasma sample from a normal donor distributed in June 1997 was 73 m/dl compared with an overall median value of 57.5 m/dl by conventional methods for vWF:Ricof (P <0.0005). This and other similar results prompted the UK NEQAS Steering Committee to recommend separate performance analyses of results obtained by users of the ELISA method compared with those obtained by other methods for vWF:RiCof determination. The difference between ELISA vWF “Activity” results and those obtained by conventional vWF:RiCof assays was further emphasised following a recent UK NEQAS distribution of a sample obtained from a plasmapheresis donor with well-characterised type 2A vWD (Val 844 Asp). The median vWF:Ag, obtained by 123 participants was 31m/dl, (range 27.5-66.5 m/dl). The median vWF “activity” level, obtained by 27 users of the vWF ELISA method was 20.0 m/dl, (range 4.556.0 m/dl), compared with a median value of 5.0 m/dl (range 0.0049.0 m/dl) obtained by 68 participants using standard techniques for vWF:RiCof (P <0.0002). Thus, the phenotypic identification of a type 2A vWD subject was readily confirmed by the majority of laboratories performing standard vWF:RiCof assays. The difference between the vWF:Ag results and those obtained with the ELISA was markedly less than that obtained by participants performing vWF:RiCof assays. Consequently, it seems unlikely that the phenotypic identification of the subject with type 2A would have been made with the same degree of confidence by users of the ELISA assay compared with those who used conventional vWF:RiCof assays. The diagnosis of vWD is not always easy. The growing popularity of a test which, in EQA surveys, has been shown to give widely differing results from the ristocetin cofactor assays that it is meant to replace, clearly demands further discussion in respect of its role and significance in the diagnosis of vWD. These issues could be appropriately addressed by the von Willebrand Subcommittee of the SSC of the ISTH.