Binding of affinity-purified anti-PF4–heparin HIT IgG to platelets during heparin-induced platelet aggregation.

Platelet-rich plasma (300 × 10⁶ platelets/mL) was mixed with HIT IgG (12 μg/mL) and ¹²⁵I-IgG tracer in a platelet aggregometer at 37°C for 12 minutes before heparin (0.5 U/mL) was added. The reaction was sampled 2 minutes before heparin (time point A), either at the start (time point B) or in the middle (time point C) of aggregation, and when platelets were fully aggregated (time point D). Samples were centrifuged through a 17% sucrose cushion to separate platelets from unbound IgG. Specific binding of patient 1 (▪) and patient 2 (•) HIT IgG was measured by using HIT¹²⁵I-IgG tracer, whereas nonspecific binding (open symbols) was determined with normal ¹²⁵I-IgG instead. The gray trace shows a typical aggregation profile that progresses from unaggregated to fully aggregated on an arbitrary scale. IgG bound is the mean ± SE of 3 to 8 experiments.