Monomeric GPVI but not dimeric GPVI binds to fibrin. SPR was performed using OneStep titration as described in "Materials and methods." (A) Representative SPR binding curve using 1 μM of GPVI. The calculated KD was 302 ± 5 nM. Solid-based binding assay was performed in nunc maxisorb 96-well plates coated overnight with 10 μM of bovine serum albumin (BSA), collagen, fibrinogen, or fibrin. (B) Monomeric (red bars) or dimeric (blue bars) GPVI (100 nM) was incubated as described. (C) Revacept, a dimeric GPVI, was allowed to bind to 10 μM of BSA, collagen, human fibrinogen, fibrin, D-dimer, or fragment E (Frag E). (D) Monomeric GPVI was allowed to bind to 10 μM of human fibrinogen, fibrin, D-dimer, or fragment E. Bound GPVI was detected using HRP coupled to an anti-6×His monoclonal antibody for monomeric GPVI or an anti-human immunoglobulin for dimeric GPVI and Revacept. The histograms (mean ± standard deviation) show the results from 5 independent experiments. **P < .01, ***P < .001 compared with a control (BSA or fibrinogen). OD, optical density.
