Fig. 2. Rac1 and Rac2 affect cytoskeletal organization in macrophages. (A) Wt, Rac2^–/– and Rac1/2^–/– BMMs in macrophage growth medium were fixed and stained with TRITC-phalloidin and anti-ß-tubulin antibodies to show F-actin and microtubules. Confocal images are shown of the basal plane of cells. (B) Quantification of F-actin levels in Wt, Rac2^–/– and Rac1/2^–/– BMMs from TRITC-phalloidin staining of paraformaldehyde-fixed cells (see Materials and Methods). Bars, 10 µm. (C) Quantification of dorsal ruffling. BMMs were stimulated with CSF-1 and stained as in (B). Cells were considered to be ruffling if more than one dorsal ruffle was present. *P<0.05 and **P<0.01 compared with levels in Wt BMMs, n=30 (Student's t-test). (D) Dorsal ruffling in response to CSF-1 stimulation in Wt, Rac2^–/– and Rac1/2^–/– BMMs. BMMs were starved of CSF-1, then stimulated with CSF-1 for 30 minutes, fixed and stained with TRITC-phalloidin to show F-actin. Confocal images were taken through the medial plane of cells to show membrane ruffles; arrows indicate membrane ruffles.