Tissue Microarray Analysis of TNBC

Tissue microarray of TNBC patients with information of clinic-pathological parameters was purchased from Outdo Biotech (HBreD090Bc01; Shanghai, China). Tissue samples were pre-stained with Ki67. All procedures were approved by the Ethical Committee of Tongji Hospital, China. Informed consent was obtained from all subjects. For immunohistochemical staining, antigen retrieval, blocking of non-specific binding and incubation of primary antibodies at 4 °C overnight were conducted sequentially. The primary antibody of anti-WDHD1 (HPA001122, Sigma-Aldrich, 1:500) was used. After incubation with secondary goat anti-rabbit immunoglobulin conjugated to peroxidase-labelled dextran polymer (SV0002; Boster) at 37 °C for 1 h, visualisation, counterstaining with haematoxylin and mounting were performed. Semi-quantitative evaluations of protein expression were scored on the basis of the intensity and the percentage of WDHD1-positive tumour cells as previously described^{59 (link)–62 (link)}.