| Abstract [eng] |
Practical Application of Diagnostic Guidelines and Implementation of New Markers of Von Willebrand Disease Background. Von Willebrand disease (vWD) is the most common genetic bleeding disorder caused by missing or defective von Willebrand factor (vWF). There are three main types of vWD based on qualitative or quantitative defects of VWF. Type 1 vWD is most common type. It causes lower-than-normal levels of VWF to occur in body. Meanwhile Type 3 vWD is very rare. People with type 3 vWD have a quantitative deficiency of vWF. People with type 2 vWD have a qualitative deficiency of vWF. Type 2 is clasified into subtypes, including: 2A, 2B, 2M, 2N. Laboratory diagnosis of vWD requires a series of assays. In this master thesis Von Willebrand factor Ristocetin Cofactor (vWF:RCo) and von Willebrand factor collagen binding (vWF:CB) assays were applied as new additional vWD markers. Methods. vWF:RCo and vWF:CB assays were implemented and evaluated by precision, measured bias, linearity over the measuring interval or analytical measurement range, limit of detection, limit of quantitation, interference, reagent carryover, reference interval or decision value. 29 blood specimens were collected from 2 groups of individuals: 20 healthy persons and 9 patients with confirmed or suspected vWD. All individuals were tested for vWF:RCo (ABP VWF Ricof Assay, ABP, UK) and vWF:CB (Asserachrom VWF:CB, Diagnostica Stago, France). Tests results were compared against a reference vWF:Ac method (Innovance VWF:Ac, Siemens Healthcare Diagnostics, Germany) to determine analytical accuracy. Additionally, patients were evaluated for other vWD markers: factor VIII, vWF antigen (vWF:Ag), platelet aggregation with ristocetin. Results. Performance of both implemented assays according to internal quality control requirements were shown to be acceptable. Evaluation by biological variation concept was acceptable for vWF:CB assay, but not for vWF:RCo. Comparison between different methods revealed that vWF:RCo does not approve raised criteria. Surrogate vWD markers (ratios of vWF:RCo and vWF:CB to vWF:Ag) have shown statistically significant correlation r=0,741 (p=0,001) and r=0,699 (p=0,011) with corresponding surrogate vWD markers of reference method (ratio of vWF:Ac to vWF:Ag). Conclusion. Only vWF:CB assay has proved it’s usefulness as an additional vWD marker, but vWD cases analysis revealed that this functional activity test is not enough for proper subtyping of vWD. Several other methods, such as vWF multimer analysis or flow cytometry, are needed. Key words. Von Willebrand disease, von Willebrand factor, von Willebrand factor Ristocetin Cofactor assay, vWF:RCo, von Willebrand factor collagen binding assay, vWF:CB. |
