Infinium methylation epic

Tumor DNA was extracted from freshly frozen tissue samples using the Qiagen DNeasy Blood & Tissue Kit (Cat NO./ID 69504) according to the manufacturer’s instructions. 500 ng of DNA were extracted from each tissue sample. DNA was sent to the Genotyping facility at the German Cancer Research Center (Heidelberg, Germany). All patient samples were analyzed using either Illumina Infinium Methylation EPIC or HumanMethylation450 BeadChip arrays according to the manufacturer’s instructions. Affiliation predictions were obtained from a DNA methylation-based classification web platform for central nervous system tumors (www.molecularneuropathology.org, version 11b4). Next, a t-SNE analysis was performed and compared with the genome-wide DNA methylation profiles from the brain tumor reference cohort [24 (link)] as well as with a previous series of ZFTA:RELA-fused EPN [3 (link)] and with the series of ZFTA-fused ependymomas reported by Zheng et al. [9 (link)]. Data was generated at the DKFZ Genomics and Proteomics Core Facility (Heidelberg, Germany) as previously described [24 (link)].

Methylation analysis was performed on DNA samples from saliva (n=450) and whole blood (n=40) using the Illumina Infinium Methylation EPIC, largely as described previously [22 (link)]. Data were adjusted for known epigenetic covariates, and surrogate variable analysis was performed via the sva inference module [23 ]. Our in-house-developed tool, CandiMeth [24 (link)], will be used to streamline the methylation analysis of gene lists of interest.

For each cohort, cord blood was collected during delivery and DNA was isolated according to standard protocols. DNA was then bisulfite-treated according to standard protocols and loaded onto Infinium HumanMethylation450 BeadChip or Infinium MethylationEPIC arrays (Illumina, San Diego, CA). Array images were scanned and raw methylation intensities were normalized and subjected to quality control according to cohort-specific procedures. For some cohorts, batch correction was applied before analysis (PREDO, INMA and Project Viva), while for the others batch variables were included as covariates in the analysis. The epigenetic data were already pre-processed within each cohort at the time of the analysis, so it was not possible to standardize the procedure across all cohorts. Previous studies in the PACE consortium used this procedure successfully [28 (link)–30 ]. Full information on the methods used within each cohort is reported in the Supplementary Methods and Materials.

Tumor DNA was extracted from freshly frozen tissue samples using the Qiagen DNeasy Blood & Tissue Kit (Cat NO./ID 69504) according to the manufacturer’s instructions. 500 ng of DNA were extracted from each tissue sample. DNA was sent to the Genotyping facility at the German Cancer Research Center (Heidelberg, Germany). All patient samples were analyzed using either Illumina Infinium Methylation EPIC or HumanMethylation450 BeadChip arrays in accordance with the manufacturer’s instructions. Affiliation predictions were obtained from a DNA methylation-based classification web platform for central nervous system tumors (https://www.molecularneuropathology.org, version 12.8). Next, a t-Distributed Stochastic Neighbor Embedding (t-SNE) analysis was performed and compared with the genome-wide DNA methylation profiles from the brain tumor reference cohort [10 (link)] and the previous series of NET, PLAGL1-fused [29 (link)]. Data were generated at the DKFZ Genomics and Proteomics Core Facility (Heidelberg, Germany) as previously described [10 (link)].