Indium tin oxide coated glass slide

FFPE blocks of ileum and jejunum from adult C57Bl/6 mice were donated by the Institute of Pathology, Technical University Dresden. A human ovarian teratoma section was provided by ProteoPath GmbH (Trier, Germany). A tissue microarray (TMA) of six different human tumors selected for being examples of their type including mantle cell lymphoma, seminoma, squamous cell carcinoma of the lung, leiomyoma, breast cancer, and melanoma was constructed from tissues collected at the Institute of Pathology, University Heidelberg, Institute of Pathology at the Technical University of Munich, and ProteoPath. Patients provided informed, signed consent and all sites collected the tissues with approval from the respective ethics committee (reference no. from the biobank of the National Cancer Centre (NCT: 2097).
All tissues were processed for paraffin embedding according to the respective automated protocols at each collection site. Tissue samples were fixed for 12–24 h in 10% neutral buffered formalin, dehydrated in graded ethanol, cleared in xylene, and embedded in molten paraffin. FFPE sections (3–5 µm) were mounted onto indium‐tin‐oxide (ITO) coated glass slides (Bruker Daltonik, Bremen, Germany) precoated with 1:1 poly‐l‐lysine (Sigma–Aldrich, Munich, Germany) and 0.1% IGEPAL CO‐630 (Sigma),18 dried overnight at 37 °C, and then stored at room temperature until MALDI–MSI analysis.

2,5-Dihydroxybenozic acid (DHB), α-cyano-4-hydroxycinnamic acid (CHCA), N,N′-dihydroxysuccinimidyl carbonate (DSC), N,N-dimethyl-amino-p-phenylenediamine, trifluoroacetic acid (TFA), 4-hydroxy-3-methoxycinnamaldehyde (CA), 2,3-diphenyl-pyranylium tetrafluoro-borate (DPP-TFB), poly-l-lysine solution, acetonitrile (ACN) and methanol (MeOH) were obtained from Sigma-Aldrich (St. Louis, MO, USA). The standards of glycine, cysteine, leucine, glutamine, tryptophan, dopamine, and norepinephrine were also purchased from Sigma-Aldrich. Indium tin oxide (ITO)-coated glass slides were purchased from Bruker Daltonics (Bremen, Germany). p-N,N,N-trimethylammonioanilyl N-hydroxysuccinimidyl carbamate iodide (TAHS) was synthesized as descried previously (Shimbo et al. 2009 (link)).

All the solvents and reagents were purchased from Sigma Aldrich (St. Louis, MO, USA). Complete mini EDTA-free Cocktail protease inhibitors were purchased from Roche. Indium tin oxide (ITO) coated glass slides were purchased from Bruker Daltonics (Bremen, Germany), polyethylene naphthalate (PEN) membrane slides (1.0 mm) were purchased from Carl Zeiss (Carl Zeiss Microsystems GmbH, Göttingen, Germany) and carboxylate modified beads, Sera-Mag 4515 and Sera-Mag 2415 SpeedBeads were purchased from Thermo Fisher Scientific.

All tissue specimens were collected intraoperatively at the Department of Gynecology, Charité—Universitätsmedizin Berlin. Written, informed consent was obtained from all patients. The study was ethically approved by the Charité Institutional Ethics Committee (ethical vote number 207/2003). EOC tissue histotyping was performed by an experienced pathologist at the Institute of Pathology, Charité—Universitätsmedizin Berlin. All surgical specimens were formalin-fixed and paraffin-embedded (FFPE) according to standard protocols. The assembly of the tissue microarray (TMA) specimens was performed at the Institute of Pathology, Charité—Universitätsmedizin Berlin. The investigated material included four TMA slides, each of which consisted of tissue cores of different EOC histotype, namely LGSOC, CCC, EC, or non-malignant BOT (tissue cores included in the analysis are shown in Supplementary Material Figure S1). Five-μm-thick TMA sections were water bath-mounted on indium-tin-oxide (ITO)-coated glass slides (Bruker Daltonics, Bremen, Germany). After overnight drying at room temperature, tissue slides were stored protected from light at 4 °C until further use.

Indium tin oxide (ITO)-coated glass slides were obtained from Bruker Daltonics (Bremen, Germany). The gold-target (purity grade > 99.995%) used for sputtering was obtained from Kurt J. Lesker Company (Hastings, England). The reagents and solvents for staining were hematoxylin and HPLC grade xylene supplied by Sigma-Aldrich (Steinheim, Germany) and ethanol (96% purity, supplied by Scharlau, Sentmenat, Spain).

Primary surgical resection specimens were preserved from gastric adenocarcinoma patients (n = 106, Table 3). Samples were collected between 1995 and 2005 at the Department of Surgery, Klinikum Rechts der Isar, Munich, Germany. Data were acquired with approval from the ethics committee of the Technical University Munich. None of the patients had received pre- or perioperative neoadjuvant treatment. Follow-up data for the patients were assessable and survival rates could be calculated using the date of last follow-up or death. The mean age at surgery was 69 years and the median survival time was 303 months. The used tissue microarray (TMA) was constructed by sampling tumor tissue with a core size of 1 mm from each paraffin-embedded tissue block. FFPE sections (3 μm) were mounted onto indium-tin-oxide (ITO)-coated glass slides (Bruker Daltonik GmbH, Bremen, Germany) pretreated with 1:1 poly-L-lysine (Sigma-Aldrich, Munich, Germany) and 0.1% Nonidet™ P-40 (Sigma-Aldrich).

Indium tin oxide (ITO)-coated glass slides were obtained from Bruker Daltonics (Bremen, Germany). The gold target used for sputtering coating was obtained from Kurt J. Lesker Company (Hastings, England) with a purity grade higher than 99.995%. HPLC grade xylene was supplied by Sigma–Aldrich (Steinheim, Germany), and ethanol (96% purity) was supplied by Scharlau (Sentmenat, Spain).