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The LN stromal cells responsible for mediating tolerance to Tyr369 have characteristics of lymphatic endothelial cells. (a) Staining of CD45neg LN stromal cells with antibodies against 10.1.1 and CD31. (b) RT-PCR for tyrosinase (Tyr) and β-actin mRNA in CD45+ cells and the indicated LN stromal cell subpopulations was evaluated as described in Fig. 1. Results shown are representative of five independent experiments. (c) Proliferation of CFSE-labeled naive Thy1.2+ FH T cells co-cultured with cells of the indicated LN stromal cell subpopulations from Thy1.1+ tyrosinase+ and albino animals was assessed as in Fig. 1. Results are from two independent experiments performed with stromal cells isolated from six pooled mice each. (d, Top) Cells with dual staining of 10.1.1 and CD31 are located in the medulla (M) and in the interfollicular areas (arrows) but not in the T cell zone (T) or B cell follicles (B; 40×). Staining is representative of multiple magnifications and fields from two independent experiments consisting of two separate LNs from two mice. Bars, 1 mm. (d, Bottom) Higher magnification (400×) of the medulla (left) and the interfollicular region (right) showing intermingling of 10.1.1+ LEC and Thy1.1+ T cells. Staining is representative of multiple magnifications and fields from two independent experiments consisting of two separate LNs from one mouse. Bars, 400 µm.