Accurate diagnosis of von Willebrand disease (VWD) depends on the quality, precision, and variability of the laboratory assays. The North American Specialized Coagulation Laboratory Association (NASCOLA) is a provider of external quality assessment (EQA) for approximately 60 specialized coagulation laboratories in North America. In this report, NASCOLA EQA data from 2010 to 2021 are reviewed for trends in methodology and precision among various assays. In particular, recent ASH ISTH NHF WFH (American Society of Hematology, International Society on Thrombosis and Haemostasis, National Hemophilia Foundation, and World Hemophilia Federation) guidelines for diagnosis of VWD are reviewed in light of EQA data. In contrast to other geographic regions, laboratories in North America predominantly use three-assay screening panels (antigen, platelet-binding activity, and factor VIII [FVIII] activity) rather than four-assay panels (antigen, platelet-binding activity, FVIII activity, and collagen-binding activity). They also use latex immunoassays rather than chemiluminescence immunoassays, and the classic ristocetin cofactor (VWF:RCo) assay and monoclonal antibody (VWF:Ab) assay to assess VWF platelet-binding activity over newer recommended assays (VWF:GPIbM and VWF:GPIbR). Factors that may be influencing these North American practice patterns include lack of Food and Drug Administration approval of the VWF:GPIbM, VWF:GPIbR, collagen binding assays, and chemiluminescence methodologies, and the influence of the 2008 National Heart, Lung, and Blood Institute guidelines on laboratory practice. Lastly, systems-based solutions are urgently needed to improve the overall accuracy of laboratory testing for VWD by minimizing preanalytical variables and adopting assay standardization.

SummaryCR1/CR2 chimeric receptors in which various short consensus repeats (SCRs) of CR1 were attached to CR2 were transiently expressed on COS cells, and assessed for the binding ofpolymerized C3b (pC3b) and anti-CR2 by immunofluorescence. Of COS cells expressing chimeras containing SCR 1-4, 1-3, 2-4, 1-2, and 2-3 of the long homologous repeats (LHRs) -B or -C, 96%, 66%, 23%, 0%, and 0%, respectively, bound pC3b. K562 cells were stably transfected with wild-type CR1, deletion mutants of CR1, and the CR1/CR2 chimeras, respectively, and assayed for binding of 12SI-pC3b. The dissociation constants (Kd) for pC3b of wild-type CR1 and the LHR-BD and -CD constructs were in the range of 1.0-2.7 nM, and of the CR1/CR2 chimeras containing SCRs 1-4, 1-3, and 2-4 of LHR-B or -C were 1.8-2.4, 6-9, and 22-36 nM, respectively. The factor I-cofactor function of the CR1/CR2 chimeras paralleled the C3b-binding function of the constructs. A CR1/immunoglobulin (Ig) chimeric protein was prepared by fusing SCRs 1-4 of LHR-B to the heavy chains of a murine F(ab')2 anti-nitrophenacetyl (NP) monoclonal antibody. The (CR1)2-F(ab')a chimera, which retained its specificity for NP, was as effective as soluble, full-length CR1 in binding pC3b, serving as a cofactor for factor I-mediated cleavage of C3b, and inhibiting activation of the alternative pathway, indicating that the bivalent expression of these SCRs reconstitutes the alternative pathway inhibitory function of CR1. The feasibility of creating CR1/Ig chimeras makes possible a new strategy of targeting complement inhibition by the use of Ig fusion partners having particular antigenic specificities.

In 2010-2012, the North American Specialized Coagulation Laboratory Association (NASCOLA) distributed 12 proficiency testing challenges to evaluate laboratory testing for protein S (PS). Results were analysed to assess the performance of PS activity, PS free antigen, and PS total antigen testing. Statistical analysis was performed on the numeric results and qualitative classification submitted for each method. There were 2,106 total results: 716 results from PS activity assays, 833 results from PS free antigen assays, and 557 results from PS total antigen assays. The three assay types performed well in the classification of five normal samples and nine abnormal samples, although certain PS activity methods were more likely to classify normal samples as abnormal and one PS total antigen assay was more likely to classify abnormal samples as normal. PS activity methods were affected by interfering substances such as heterozygous or homozygous factor V Leiden mutation (underestimation) and the anticoagulant drug rivaroxaban (overestimation). In conclusion, NASCOLA laboratories using a variety of PS assays performed well in the classification of clearly normal and abnormal samples. Laboratories performing PS activity assays should be aware of potential interferences in samples positive for FV Leiden or containing certain anticoagulant medications.

Background Laboratory diagnosis of von Willebrand Disease (VWD) is complex. Reliance on laboratory testing can be problematic as different VWD screening panels, assays and methodologies can produce analytic variability in test results. Objectives To compare the degree of imprecision among the VWD assays and within the platelet binding activity (PBA) assays, to determine the consensus among the VWD assays for correct classification of sample results, and to determine consensus among laboratories’ von Willebrand factor (VWF) multimer interpretations and final interpretations of the VWD panels. Patients/Methods Proficiency testing results from the North American Specialized Coagulation Laboratory Association (NASCOLA) submitted by laboratories from 2010 to 2019 for all normal, type (T) 1 VWD and T2 VWD samples were analysed. Results and Conclusions Imprecision was lowest for VWF antigen and highest for collagen binding activity (CBA) with median coefficient of variation (CV) of 12% (interquartile range (IQR) 7%) and 23% (IQR 21%) respectively. Within the VWF PBA assays, the gain‐of‐function mutant GP1b binding (VWF: GP1bM) methods had the least imprecision (CV 9%, IQR 10%). All assays, including the various PBA methods had excellent consensus. The majority of laboratories agreed that normal (median consensus‐82%, IQR 16%) and T1 VWD (median consensus‐100%, IQR 9%) samples had normal multimer distribution. Consensus among laboratories for final interpretations was excellent for normal samples (median 81%, IQR 8%), good for T1 VWD samples (median 59%, IQR 9%), and fair for T2 VWD samples (median 44%, IQR 21%). Consensus on final interpretation decreased as sample complexity increased.