CD163 is considered to be a macrophage-specific marker and is generally not expressed in cell types other than monocytes/macrophages [26 (link)]. Based on the cell fusion theory, we used CD163-expression as a surrogate marker for macrophage phenotype in breast cancer cells [16 (link), 27 (link)]. Five-micrometer sections were obtained from formalin-fixed paraffin-embedded TMA tumor specimens. The sections were de-paraffinized in xylene and hydrated in a series of graded alcohols, pretreated with heat-induced epitope retrieval and trisethylenediaminetetraacetic acid buffer (1 mM, pH 9, 20/5/20 min; Decloaking Chamber NxGen, Biocare Medical), and stained for CD163 (anti-human, monoclonal antibody, clone 10D6, Novocastra, Leica). Staining for estrogen receptor (ER; clone SP1, Ventana Roche), progesterone receptor (PR; clone 1E2, Ventana Roche), Ki-67 (clone MIB-1, Dako Agilent), and human epidermal growth factor receptor 2 (HER2; clone 4B5, Ventana Roche) was done according to clinical laboratory standards. All slides were scanned to digital images using the Hamamtsu NanoZoomer XL (Visiopharm LRI AB). Evaluation of immunostaining was performed using ImageScope viewing software (Leica Biosystems).
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