Src and ezrin expression in human breast cancer. (A) IHC staining of breast normal and tumour tissue was used to validate Src and ezrin antibodies. Scale bars = 75 μm. (B) Cy5 IF staining was used to assess the expression levels of Src and ezrin in the 63-patient breast cancer TMA. Representative tumour cores with high and low expression of Src and ezrin are presented. Images were acquired using Aperio ScanScopeFL. Scale bars = 100 μm. (C) Association between dichotomized ezrin AQUA scores (cutoff: median) and continuous Src AQUA scores based on IF-stained 63-TMA. (D, E) Continuous AQUA scores of total Src and ezrin were correlated against dichotomized LVI status. Statistical associations were assessed using the unpaired Student's t test. Associations were considered significant when P <0.05. AQUA: automated quantification analysis IF: immunofluorescence; IHC: immunohistochemistry; LVI: lymphovascular involvement; TMA: tissue microassay.