Activation-independent antibody-induced depletion of GPVI from human platelets circulating in NOD/SCID mice. (A) Washed human platelets resuspended in Tyrode-HEPES buffer at 3.0 × 108 platelets/mL were incubated for 2 minutes in the presence of 10 μg/mL isotype control mAb (left) or the mAb 6B12 (right). Note that 6B12 IgG neither induces nor blocks platelet aggregation. Platelets were fully reactive, as shown by later addition of CRP. (B) Concentrated human PRP (200 μL) containing 2 × 109 platelets/mL was injected into the retro-orbital plexus of a NOD/SCID mouse, followed by an intraperitoneal injection of 30 μg indicated Fab. Platelets in whole blood drawn from the tail vein at 0, 0.5, 3, and 24 hours after injection were assessed for surface expression of human GPIIb-IIIa and GPVI. Note that GPVI surface expression is not noticeably affected on platelets from mice treated with anti–PECAM-1 Fabs (top 4 panels) but decreases significantly in mice treated with the GPVI-specific Fab, 6B12.
