Immunolocalization of Rac1, PAK, and cortactin in thrombin-stimulated platelets.

Resting or stimulated platelets (0.5 UI/mL thrombin for 1 minute at 37°C) were fixed and labeled with 1-10 μg/mL anti-Rac1 MoAb, anti-PAK1/2 rabbit polyclonal antibody, or anti-cortactin monoclonal antibody for 30 minutes at 4°C. After washes in PBS (pH 7.40), samples were labeled with either secondary goat antimouse antibodies coupled to FITC or secondary goat antirabbit antibodies coupled to rhodamine. Samples were analyzed by confocal fluorescence microscopy (×100). Controls with irrelevant mouse or rabbit IgGs are shown in (C). Results are representative of 3 separate experiments.