Crucial Contributions by T Lymphocytes (Effector, Regulatory, and Checkpoint Inhibitor) and Cytokines (TH1, TH2, and TH17) to a Pathological Complete Response Induced by Neoadjuvant Chemotherapy in Women with Breast Cancer

Figure 5

IL-1 (a, b), IL-2 (c, d), and IFN-γ (e, f) expression in the sections of LLABCs, 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 IL-1 (Abcam, ab8320) at a 1 : 150 dilution overnight at 4°C and MAbs to IL-2 (Abcam, ab92381) at a 1 : 500 dilution for 30 mins at RT and polyclonal Abs to IFN-γ (Abcam, ab9657) at a concentration of 4 μg/mL for 30 mins at RT, respectively. 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, e) Low level of expression; (b, d, f) high level of expression. The score (% of positive cells (brown membrane/cytoplasmic-stained tumour and immune cells) × intensity of staining (1 to 3)) was used to assess the level of expression; low was ≤100 and high was >100. Scoring performed on whole tissue section (>10 HPFs); Tu: tumour and Ly: lymphocyte.

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