Patient anti–β₂ GPI antibodies interfere with the interaction between VWF and β₂ GPI. (A) Total aggregation of washed platelets induced with 2 μg/mL collagen was set at 100%. Relative aggregation in the presence of β₂ GPI (2.2 μM) and in the presence of β₂ GPI and patient IgG of 3 different patients (100 μg/mL) was calculated. All aggregation experiments were performed with platelets of 5 different blood donors. Data represent the mean (± SD) of the effect of antibodies isolated from 1 patient and are representative for the data found with the IgGs from the other 2 patients. *P = .02, ***P < .001. (B) β₂ GPI–coated wells were incubated with biotinylated VWF/A1-R1306Q (1.1 μg/mL) in the presence of different concentrations of affinity-purified antiprothrombin antibodies (▩) or anti–β₂ GPI antibodies (□). Bound VWF/A1-R1306Q was detected using streptavidin-HRP. Experiments were performed with antibodies purified from plasma of 3 different patients, and a typical example is presented. Data show the mean (± SD) of 3 experiments. (C) β₂ GPI–coated wells were incubated with plasma of healthy individuals (n = 16), plasma of patients negative for LAC activity (n = 10), plasma of patients positive for LAC activity not due to anti–β₂ GPI antibodies (n = 10), or plasma of patients positive for LAC activity due to anti–β₂ GPI antibodies (n = 10) for 2 hours at 37°C. After washing, wells were incubated with biotinylated VWF/A1-R1306Q (1.1 μg/mL) for 30 minutes at 37°C. Bound VWF was detected with streptavidin-HRP. Residual binding determined in 3 independent experiments is presented in a box-and-whiskers plot. ***P < .001.