Analysis of tumor-infiltrating cells in human and mouse tumors. (A) Tumor sections from patients with breast cancer were stained for the macrophage marker CD68 (brown) and counterstained with hematoxylin. (B-I) Sections of subcutaneous MMC tumors in neu-tg mice. Tumors were analyzed 3 weeks after MMC transplantation. (B) Adherens junction marker E-cadherin (green) and stroma marker laminin (red); (C) endothelial marker CD31 (green) and laminin (red); (D) panleukocyte marker CD45 (green) and laminin (red); (E) CD45 (green) and relaxin receptor (red); (F) macrophage marker F4/80 (green) and laminin (red); (G) CD4 T-cell marker (green) and laminin (red); (H) CD8 T-cell marker (green) and laminin (red); (I) NK cell marker NK1.1 (green) and laminin (red). Nuclei are stained with DAPI (blue). The scale bar represents 100 μm. (J) Quantification of immune cells in tumor-draining lymph nodes. Intracellular cytokine staining for IFNγ and CD4, using anti–IFNγ-PE antibodies and anti-CD8, anti–CD4-FITC antibodies. (K) Quantification of immune cells in tumor-draining lymph nodes. Frequency of Neu-specific CD8⁺ cells in splenocytes was determined by Tetramer assay.²⁶  Characteristic samples are shown. The numbers in the quadrants represent the precentage of positive cells.