Virtuoso v 5

IHC was performed following standard procedures. Briefly, tumor tissues were extracted, fixed in formalin, paraffin-embedded and subsequently sectioned in 5 µm coronal sections with a Leica rotary microtome. The following primary antibodies were used: Ki67 (Roche; Ref 790-4286, dilution 1:100), p53 (Roche; Ref 790-2912, predilution 1:20), IDH1 (vitro master diagnostica; Ref R132H antibody H09, predilution) and ATRX (Sigma-Aldrich; Ref HPA001906, 1:200). IHCs were analyzed by a senior pathologist of the Service and were performed following the manufacturer’s instructions on the Roche Ventana Benchmark ULTRA System with ethylenediaminetetraacetic acid (EDTA) pH 8.5 antigen retrieval. Sections were scanned with Virtuoso v.5.6.1 software (Ventana Medical Systems, Roche).

IHC was performed following standard procedures. Briefly, whole brains were extracted, fixed in formalin, paraffin‐embedded, and subsequently sectioned in 5 µm coronal sections with a Leica rotary microtome. Sections were taken at lateral geniculate nucleus of the thalamus in the temporal lobe, which was considered our topographic reference point. All sections included hippocampus, temporal horn, and entorhinal cortex. The following primary antibodies were used: CD3 (Roche, Ref.: 790‐4341, Clone: 2GV6), CD4 (Roche, Ref.: 790‐4423, Clone: SP35), CD8 (Roche, Ref.: 790‐4460, Clone: SP57), CD68 (Roche, Ref.: 790‐2931, Clone: KP‐1), IBA1 (Wako; Ref.: 019‐19741), GFAP (Roche, Ref.: 760‐4345), Interferon‐γ (Abcam, Ref.: ab9657), IL1α (Abcam, Ref.: ab9614), SOX2 (Cell Marque™, Ref.: 760‐4621, Clone: SP76), and p16^INK4a (Roche, Ref.: 805‐4713). IHC was performed following the manufacturer´s instructions on the Roche Ventana Benchmark ULTRA System with ethylenediaminetetraacetic acid (EDTA) pH 8.5 antigen retrieval. Hematoxylin–eosin staining of SVZ sections was performed using standard procedures. Sections were visualized with a light microscope using the 10×, 20×, and 40× objective and then scanned with Virtuoso v.5.6.1 software (Ventana Medical Systems, Roche).

Tissue slides for immunohistochemistry (IHC) were obtained as follows: Briefly, lung lobes from autopsies were extracted, fixed in formalin, paraffin-embedded and subsequently sectioned in 4 μm sections with a Leica rotary microtome. Sections from peripheral and central regions of right and left lung lobes were obtained for further analysis. In the case of lung samples coming from biopsies, the same procedure was followed with the specific sample obtained. IHC was performed following the manufacturer's instructions on the Roche Ventana Benchmark ULTRA System with ethylenediaminetetraacetic acid (EDTA) pH 8.5 antigen retrieval. The following primary antibodies were used: SOX2 (Roche Ventana, cat number: 760-4621), p63 (Roche Ventana, cat number: 790-4509), KRT5 (Roche Ventana, cat number: 790-4554), TTF-1 (Roche Ventana, cat number: 790-4756), p16^INK4A (Roche Ventana, cat number: 805-4713), p21^CIP (Roche Ventana, cat number: 760-4453), Lamin B1 (Cell Signaling Technology, cat number: 12586S; dilution 1:300), Ki67 (Roche Ventana, cat number: 790-4286) and KRT14 (Roche Ventana, cat number: 760-4805). Hematoxylin-eosin was performed using standard procedures. Sections were visualized and scanned with Virtuoso v.5.6.1 software (Ventana Medical Systems, Roche).