American Society of Hematology

Fig. 5.

$Fig. 5. Immunohistologic analysis of JAM-1, -2, and -3 expression in murine lymphoid organs. / (A) Frozen sections of murine mesenteric lymph nodes were stained for JAM-1, JAM-2, JAM-3, MAdCAm, or PECAM using peroxidase-coupled secondary antibodies. The negative control was obtained using an IgG fraction from nonimmune rabbit serum, followed by antirabbit coupled to peroxidase. Peroxidase-stained structures appear brown, and the hemalum counterstain of tissue appears blue. Germinal centers (GC), lymphatic sinuses (LS), and HEVs (arrowheads) are indicated. Magnification × 40. (B) Frozen sections of murine mesenteric lymph nodes were triple stained for JAM-1, JAM-2, and JAM-3 as indicated. Single-color images were acquired using specific excitation and emission filter sets, and pseudocolored images were acquired using Adobe Photoshop 5.5. The absence of fluorescence leakage in the different channels was checked by single staining using the same experimental setup. Magnification × 400. (C) Differential localization of JAM-1 and JAM-2 on lymphatic sinuses. Lymphatic sinuses indicated by arrows express JAM-1 diffusely, whereas JAM-2 is concentrated in cell-cell contacts. Arrowheads indicate a vascular structure expressing JAM-1 and JAM-2. Secondary antibodies were antirat Texas Red and streptavidin-FITC to visualize, respectively, anti–JAM-2 and biotinylated anti–JAM-1 reactivities. In the merged picture, JAM-1 appears in green and JAM-2 in red. Magnification × 160.$

Immunohistologic analysis of JAM-1, -2, and -3 expression in murine lymphoid organs.

(A) Frozen sections of murine mesenteric lymph nodes were stained for JAM-1, JAM-2, JAM-3, MAdCAm, or PECAM using peroxidase-coupled secondary antibodies. The negative control was obtained using an IgG fraction from nonimmune rabbit serum, followed by antirabbit coupled to peroxidase. Peroxidase-stained structures appear brown, and the hemalum counterstain of tissue appears blue. Germinal centers (GC), lymphatic sinuses (LS), and HEVs (arrowheads) are indicated. Magnification × 40. (B) Frozen sections of murine mesenteric lymph nodes were triple stained for JAM-1, JAM-2, and JAM-3 as indicated. Single-color images were acquired using specific excitation and emission filter sets, and pseudocolored images were acquired using Adobe Photoshop 5.5. The absence of fluorescence leakage in the different channels was checked by single staining using the same experimental setup. Magnification × 400. (C) Differential localization of JAM-1 and JAM-2 on lymphatic sinuses. Lymphatic sinuses indicated by arrows express JAM-1 diffusely, whereas JAM-2 is concentrated in cell-cell contacts. Arrowheads indicate a vascular structure expressing JAM-1 and JAM-2. Secondary antibodies were antirat Texas Red and streptavidin-FITC to visualize, respectively, anti–JAM-2 and biotinylated anti–JAM-1 reactivities. In the merged picture, JAM-1 appears in green and JAM-2 in red. Magnification × 160.

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