Leica aperio at2 slide scanner

We retrieved the FFPE case study population from the Pathology Unit’s archive of the University of Naples “Federico II.” OSCC FFPE tumor samples, from surgical resections, were used to build tissue microarrays (TMAs). Three TMAs were built containing 111 tumor samples, selecting the most representative areas from each selected paraffin block, at least in duplicate. Then, 3 mm tissue cores were punched from morphologically representative tissue areas of each donor block and placed into one recipient paraffin block (3 × 2.5 cm) using a semi-automated tissue arrayer (Galileo TMA, Milan, Italy). One section of each TMA (4 μm) was stained with hematoxylin and eosin (H&E) to check the adequacy of cores.
H&E-stained and IHC-stained glass slides were digitalized at 40× using the Leica Aperio AT2 slide scanner (Leica Biosystems, Vista, CA, USA).
The study was performed according to the guidelines of the institutional ethics committee, which, in agreement with Italian law, concerning the topics of the current research, and according to Declaration of Helsinki, requires, for studies based only on retrospective analyses on routine archival FFPE-tissue, a written informed consent from the living patients, following the indication of Italian DLgs No. 196/03 (Codex on Privacy), as modified by the UE 2016/679 law of the European Parliament and Commission at the time of surgery.

H&E-stained and IHC-stained glass slides were digitalized at 40× using the Leica Aperio AT2 slide scanner (Leica Biosystems, Wetzlar, Germany) [23 (link)]. WSI images in .svs file format were analyzed with the QuPath platform. Classification was performed applying a Random Tree classifier [24 (link)]. The staining vector signal intensity was assessed and quantified to obtain a H-Score for NRP-1 tissue expression in both OSCC and OPSCC samples. NRP-1 H-Score values were categorized for each tumor site (OSCC and OPSCC) into low and high-expression groups; the threshold for categorization was selected via ROC curve analysis for the OS (Overall Survival) outcome (Figure 1). For TCGA dataset analysis, data were retrieved from the TCGA website, and mRNA levels of the NRP1 gene were analyzed and thresholded using kmplot.com analysis tools [25 (link)] (Last access on 14 July 2021).

The TMA was evaluated for presence of the tumor tissue. Only TMA cases with at least one representative core were included for further analyses. The immunohistochemistry results were first digitalised using Leica Aperio AT2 slide scanner (Leica Biosystems, Buffalo Grove, IL, USA) and then evaluated with Aperio ImageScope software (Leica Biosystems, Buffalo Grove, IL, USA). The assessment of immunohistochemistry was performed by an experienced neuropathologist (JS). Percentage of positive cells and most prevalent staining intensity (1– weak, 2– moderate, 3– strong) was noted. Average percentage of positive cells and modified H score (mHS = percentage * intensity) was used for the analysis as reported previously [18 ]. Whole sections were analyzed using the same approach.