Thrombus formation in blood from mice genetically modified to express different levels of GPVI. (A) Blood from control mice, FcR γ-chain^(+/–) mice with a 50% reduction in GPVI levels, FcR γ-chain^(–/–) mice deficient in GPVI, and FcR γ-chain^(–/–) mice that express an FcR γ-chain ITAM (ITAM PM) mutant transgene at a level that is approximately 40% of the wild type was flowed over a collagen-coated surface at 800 s^–1 Thrombus formation was abolished in the FcR γ-chain^(–/–) mice and in blood from the ITAM PM mice, although there was some adherence of single platelets. Quantitation of percent of surface coverage of the collagen-coated microslide using Image-Pro plus software demonstrates control platelets giving 70 ± 4% coverage compared with 72 ± 5% for the heterozygotes, 7 ± 1% for FcR γ-chain–deficient platelets, and 8 ± 1% ITAM PM platelets. These results are presented as ± SEM and are representative of 3 to 8 experiments. Original magnification, × 630. (B) Blood from control mice and FcR γ-chain^(+/–) mice, with a 50% reduction in GPVI levels, was flowed over differing concentrations of collagen-coated surface as indicated. Platelets adherent to the collagen-coated surface were lysed in 1% NP-40 lysis buffer and subjected to sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to polyvinylidene fluoride (PVDF) membrane. The membrane was probed with an anti-Actin antibody. These results are representative of 3 experiments. Thrombus formation varied by less than 10% as measured by protein estimation of adherent platelets between control and FcR γ-chain^(+/–) mice. (C) Blood from genetically modified mice as indicated was flowed over a collagen-coated surface at 800 s^–1 and 1500 s^–1. Platelets adherent to the collagen-coated surface were lysed in 1% NP-40 lysis buffer and subjected to SDS-PAGE and transferred to PVDF membrane. The membrane was probed with an anti-Actin antibody. These results are representative of 3 to 8 experiments.