Figure 2.

Immunohistochemistry and immunofluoresence for TIM-2 on spleen sections. (A) The anti–TIM-2 antiserum binds specifically to clusters of cells in splenic follicles. Paraffin-embedded sections from unimmunized mice were stained with an isotype-matched control rabbit antiserum (control IgG), anti–TIM-2 antiserum (TIM-2); or anti–TIM-2 antiserum blocked with an excess of the immunizing peptide (TIM-2 + peptide), followed by goat anti–rabbit IgG conjugated to horseradish peroxidase. The slides were counterstained with hematoxylin. (B) TIM-2–positive cells are predominantly B cells. Adjacent cryostat sections from unimmunized mice were co-stained for TIM-2 and B220 (top row) or TIM-2 and CD3 (bottom row). TIM-2 staining appears green (left columns), B220 and CD3 staining appear red (middle columns), and merged images are shown in the far right columns. Bars, 100 μm. (C) TIM-2–positive cells are localized to GCs. Adjacent cryostat sections from the spleen of a mouse that was immunized with SRBCs was stained for peanut agglutinin (PNA, blue, left panel) to show the GCs, and with IgD (brown, right panel) to show the surrounding follicular mantle zone. The section in the right panel was stained for TIM-2 (blue) and IgD (brown).