Figure 2.

CD68⁺ (a, b) and CD163⁺ (c, d) macrophages in the sections of axillary lymph nodes (ALNs), using IHC staining, at 400x magnification. Briefly, heat-mediated antigen retrieval was performed using citrate buffer, pH 6 (20 mins). The sections were then incubated with MAbs to CD68 (Abcam, ab955) at a 1 : 300 dilution for 30 mins at RT, MAbs to CD163 (Abcam, ab74604) at a prediluted concentration for 30 mins at RT. Polymeric HRP-linker antibody conjugate was used as secondary antibody. DAB chromogen was used to visualize the staining. The sections were counterstained with haematoxylin. (a, c) Low percentage of CD68⁺ and CD163⁺ macrophages; (b, d) high percentage of CD68⁺ and CD163⁺macrophages. The positive brown membrane-stained cells in tumour-free medullary areas of ALNs were quantified as the average % of all cells (5 HPFs).