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Histological analysis

TI Tamara Isermann
ÖŞ Özge Çiçek Şener
AS Adrian Stender
LK Luisa Klemke
NW Nadine Winkler
AN Albrecht Neesse
JL Jinyu Li
FW Florian Wegwitz
UM Ute M. Moll
RS Ramona Schulz-Heddergott
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Standardized immunohistochemical staining were performed on murine FFPE tissues. The following primary antibodies were used and listed in detail in Supplementary Table 1: p53 FL393 (Santa Cruz); cyclinD1, pan-Cytokeratin, and α-smooth muscle actin/SMA (all Abcam); phospho-HSF1 (Bioss); and pRB and Hsp70 (both Cell Signaling). The ImmPRESS™ Peroxidase polymer reagent based on 3, 3-diaminobenzidine (DAB, Vectorlabs), or Alexa Fluor®488-coupled and Alexa Fluor®647-coupled secondary antibodies (immunofluorescence) were used as detection systems. Hematoxylin (DAB) or DAPI (immunofluorescence) were used as counterstains. Stained sections were analyzed by microscopy (AxioScope, Zeiss) with ZENblue software V3.0 (Zeiss). Grading for phospho-HSF1 and p53 was as follows: p53high, >25% with intense nuclear staining; Hsp70high, >50% with high intensity.

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