IntroductionLymph nodes are one of the main sites where an effective immune response develops. Normally, axillary nodes are the first place where breast cancer produces metastases. Several studies have demonstrated the importance of immune cells, especially dendritic cells, in the evolution of breast cancer. The goal of the project is to identify differences in the patterns of immune infiltrates, with particular emphasis on dendritic cells, in tumour and axillary node biopsies between patients with and without metastases in the axillary nodes at the time of diagnosis. It is expected that these differences will be able to explain differences in survival, relapse and clinicopathological variables between the two groups.Methods and analysisThe study will involve 100 patients diagnosed with invasive breast cancer between 2000 and 2007, 50% of whom have metastases in the axillary lymph node at diagnosis. In selected patients, two cylinders from biopsies of representative areas of tumour and axillary nodes (with and without metastasis) will be selected and organised in tissue microarrays. Samples will be stained using immunohistochemical techniques for different markers of immune response and dendritic cells. Two images of each cylinder will be captured under standardised conditions for each marker. Each marker will be quantified automatically by digital image procedures using Image-Pro Plus and Image-J software. Associations of survival, relapse and other clinicopathological variables with the automatically quantified levels of immune infiltrates in patients with and without axillary node metastasis will be sought.Ethics and disseminationThe present project has been approved by the Clinical Research Ethics Committee of the Hospital Universitari Joan XXIII (Ref: 22p/2011). Those patients whose biopsies and clinical data are to be used will give their signed informed consent. Results will be published in peer-reviewed journals.
BackgroundDigital image (DI) analysis avoids visual subjectivity in interpreting immunohistochemical stains and provides more reproducible results. An automated procedure consisting of two variant methods for quantifying the cytokeratin-19 (CK19) marker in breast cancer tissues is presented.MethodsThe first method (A) excludes the holes inside selected CK19 stained areas, and the second (B) includes them. 93 DIs scanned from complete cylinders of tissue microarrays were evaluated visually by two pathologists and by the automated procedures.Results and conclusionsThere was good concordance between the two automated methods, both of which tended to identify a smaller CK19-positive area than did the pathologists. The results obtained with method B were more similar to those of the pathologists; probably because it takes into account the entire positive tumoural area, including the holes. However, the pathologists overestimated the positive area of CK19. Further studies are needed to confirm the utility of this automated procedure in prognostic studies.
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