Measurement of von Willebrand factor antigen in plasma and platelets with an enzyme-linked immunosorbent assay based on two murine monoclonal antibodies.

Two murine monoclonal antibodies, raised against von Willebrand factor (vWF), were used to construct an enzyme-linked immunosorbent assay (ELISA), for quantitation of vWF antigen (vWFAg) in human plasma and platelets. This assay had a lower limit of sensitivity of 0.0001 IU/ml in buffer, and thus is one to two orders of magnitude more sensitive than other ELISA assays which have been reported. The intraassay, interassay and interdilution coefficients of variation were 4.1, 10.4 and 9.9%, respectively. In normal plasma (n = 20), the vWFAg level was 0.83 (range: 0.42-1.25) IU/ml. In normal washed platelets (n = 10), 0.35 (0.25-0.49) IU/10(9) platelets was found. In plasma obtained from various patient groups the following vWFAg levels (geometric mean and range) were observed: von Willebrand's disease (n = 19): 0.18 (0.02-0.77) IU/ml; patients with liver cirrhosis (n = 20): 3.73 (1.68-9.20) IU/ml; patients with pregnancy-induced hypertension (n = 20): 4.14 (2.28-7.44) IU/ml and patients with malignant disease (n = 10), 2.54 (1.51-5.60) IU/ml. A linear correlation was found between vWFAg levels measured with a polyclonal antibody based Laurell electroimmunoassay (r = 0.92, n = 58) or with a polyclonal antibody based ELISA (r = 0.94, n = 64). The present assay is based on stable and reproducible reagents and allows the specific measurement of vWFAg in plasma and in platelets. This assay may constitute a useful tool for the further investigation of clinical conditions associated with changes in vWFAg levels. In addition, its high sensitivity may facilitate a more detailed study of platelet vWFAg in normal and in pathological conditions.

Prolonged bleeding time due to mitotane therapy.

After finding prolonged bleeding times in 2 patients treated with mitotane, we prospectively studied 7 patients with adrenocortical cancer on mitotane therapy. Before and 1 and 2 or more weeks after starting mitotane we determined the platelet counts, bleeding times and global coagulation parameters. All patients had a normal bleeding time before treatment. In 6 cases the bleeding time became prolonged (245-555 s). 4 patients exhibited platelet aggregation responses compatible with an aspirin-like defect. It is concluded that mitotane may cause a clinically relevant defect of platelet function.

An infrequent DNA polymorphism associated with severe von Willebrand's disease.

Genomic DNA of six unrelated Dutch patients with severe von Willebrand's disease (vWD) was submitted to restriction fragment length polymorphism analysis. We observed a strong association between a 36 kb allele detected by a partial complementary DNA probe (pvWF 1100) and the restriction enzyme XbaI with severe von Willebrand's disease. This 36 kb allele is rare (allele frequency of 7%) both in the general population and in patients with autosomal dominant types of von Willebrand's disease. Three of our six patients were found to be homozygous for this allele while two others were heterozygous. The association of this rare XbaI allele with severe vWD enables carrier detection and prenatal diagnosis in these families. The high frequency (67%) of the 36 kb allele observed in this patient group raises the possibility that a subgroup of patients with severe vWD has a genetic defect with a common origin.

Bleeding time, blood groups and von Willebrand factor.

The bleeding time in healthy volunteers was determined according to both the Ivy and the Simplate II techniques. A significantly longer bleeding time in people with blood group O than in people with non-O blood groups was demonstrated with both techniques. This difference could not be attributed to a difference in sex ratio, platelet count or haematocrit. The mean level of von Willebrand factor in blood group O is lower than in non-O blood groups, but we found no association between the level of von Willebrand factor and the bleeding time, despite the very broad range of von Willebrand factor levels in the subjects examined.

A randomized and blinded comparison of the sensitivity and the reproducibility of the Ivy and Simplate II bleeding time techniques.

The authors compared the sensitivity and the reproducibility of the bleeding time techniques according to Ivy and Simplate II. The sensitivity was studied in two groups: one group of 64 healthy volunteers and another group of 40 patients with various disorders of hemostasis, including 28 patients with Von Willebrand's disease. Ivy and Simplate II bleeding times were performed on each subject. The reproducibility was studied in 48 patients with mildly or moderately prolonged bleeding times that resulted from various disorders who had a duplicate Ivy or a duplicate Simplate II bleeding time. All subjects were randomized over the technologists and they were blinded for each other's results. By a receiver operating characteristic analysis, the Ivy method appeared to offer greater overall detection efficacy than the Simplate II method. For the Ivy method, the standard deviation of the ratios of the duplicate bleeding times was 0.37 and for the Simplate II method it was 0.33. The authors conclude that the Simplate II method is not superior in sensitivity or reproducibility to the Ivy method, which is cheaper, takes less time, and does not leave scars.

Arthropathy in von Willebrand's disease.

A 62-year-old woman with severe von Willebrand's disease and a long history of joint complaints is presented. Her history, the progressive radiological findings, the demonstration of haemarthrosis and a literature review support the view that some patients with von Willebrand's disease can suffer from an incapacitating arthropathy akin to that seen in haemophilia.