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Fig. 3 | BMC Cancer

Fig. 3

From: Tumour-draining axillary lymph nodes in patients with large and locally advanced breast cancers undergoing neoadjuvant chemotherapy (NAC): the crucial contribution of immune cells (effector, regulatory) and cytokines (Th1, Th2) to immune-mediated tumour cell death induced by NAC

Fig. 3

CD8+ T cells (a, b), FOXP3+ Tregs (c,dD) and CD56+ NK cells (E, F) in the sections of axillary lymph nodes (ALNs), using IHC staining, at 400× magnification. Briefly, heat-mediated antigen retrieval was performed using citrate buffer pH 6 (20 mins). The sections were then incubated with MAbs to CD8 (Dako, M7103) at a 1:100 dilution for 30 mins at RT, MAbs to FOXP3 (Abcam, ab20034) at a concentration of 20 μg/ml for 30 mins at RT, MAbs to CD56 (Dako, M7304) at a 1:50 dilution 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, e: low percentage of CD8+ T cells, FOXP3+ Tregs and low number of CD56+ NK cells respectively; B, d, d: high percentage of CD8+ T cells, FOXP3+ Tregs and high number of CD56+ NK cells respectively. The positively brown membrane-stained cells (CD8+ T cells) and brown nuclear-stained cells (FOXP3+ Tregs) in non-metastatic paracortical areas of ALNs were quantified as the average % of all cells (5 HPFs). CD56+ NK cells were quantified as average number of cell count per HPF in non-metastatic para-cortical areas of ALNs with the greatest accumulation of the positively brown membrane-stained cells

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