Expression of a mutant form of SHPS-1 that lacks most of the cytoplasmic region in splenocytes. (A) Frozen spleen sections from WT or SHPS-1 mutant (KO) mice at 6 weeks of age were stained both with the P84 mAb to SHPS-1 (red) and with a mAb to F4/80 (green). The SHPS-1 and F4/80 images are also shown merged. Scale bar, 100 μm. Images were visualized using an AX-70 microscope equipped with a 10×/0.4 objective lens (Olympus, Tokyo, Japan) and a PXL 1400 camera (Photometrics, Tucson, AZ). Images were analyzed with IPLab Spectrum software (Scanalytics, Fairfax, VA) and were processed with Adobe Photoshop 8.0 software (Adobe Systems, San Jose, CA). (B) Splenocytes prepared from WT and SHPS-1 mutant mice at 6 weeks of age were stained with a PE-conjugated mAb to F4/80 and biotinylated mAb P84 (thick trace) or with isotype control mAbs (thin trace) and were then incubated with FITC-conjugated streptavidin. The expression of SHPS-1 on F4/80-positive splenic macrophages was then examined by 2-color flow cytometric analysis. All results are representative of 3 separate experiments.