VWF reactivity in SCD. (A) VWF multimer distribution. The VWF multimer distribution was examined in plasma from either sickle cell patients or pooled normal plasma by electrophoresis followed by Western blotting. For each of the sickle cell patients, 0.5 μL of plasma was loaded. For pooled normal plasma, PNP I and PNP II, 1 μL and 0.5 μL of plasma was loaded, respectively. The samples were labeled according to the patient identification number and grouped by genotype. The dashed line is drawn slightly above the highest multimer in pooled normal plasma. The multimers above the dashed line in samples of sickle cell patients are considered ULVWF. (B) VWF parameters. Data from different patients are presented in different colors; patients 1, 2, 3, 10, and 11 provided multiple samples. The values expressed are relative to those obtained with pooled normal plasma, which was arbitrarily assigned a value of 1, as indicated by the dashed lines in each panel. The assays were performed independently 2 or 3 times per sample; average values are shown. VWF antigen was measured by sandwich ELISA using a polyclonal VWF antibody as the capture antibody, and the captured VWF was detected with a horseradish peroxidase–conjugated polyclonal VWF antibody. VWF:AF was determined using AU/VWFa-11 as the capture antibody in ELISA, and the bound VWF was detected by a horseradish peroxidase–conjugated polyclonal VWF antibody. VWF:TA in the plasma was determined by multiplying the VWF antigen concentration by the VWF-AF. (C) Correlation between VWF parameters and LDH. The association between LDH and each of the VWF variables was assessed by fitting mixed-effects regression models. The estimated regression line has been superimposed on the corresponding graph. The number of patients was 13, and the number of observations was 19. LDH concentration correlated significantly with VWF:AF (P = .003) and VWF:TA (P = .007) but not with VWF antigen concentration (P = .115). Patients with different genotypes are identified by different colors: black represents SS; blue, Sβ⁰; red, Sβ⁺; and green, SC. (D) VWF:TA correlates best with LDH. When the regression lines for VWF antigen, VWF:AF, and VWF:TA were plotted on the same scale, the slope of the regression line between VWF:TA and LDH was the steepest, indicating that VWF:TA is more sensitive to changes in LDH than VWF:Ag or VWF:AF.