At delivery, clinical staff were prompted to collect cord blood samples using paper or electronic “flags” on mothers’ charts. Clinicians collected cord blood from the umbilical vein using a syringe and needle. Blood samples were collected from children at the early and mid-childhood visits. Within 24 h of collection, cord blood and childhood blood samples were centrifuged at 1700 × g for 10 min at 4 °C to separate plasma, red blood cells (RBCs), and nucleated cells used for measurement of DNAm (leukocytes and nRBCs in cord blood, and leukocytes in childhood blood).
DNAm was measured in all participants with proper consent for genetic and epigenetic analyses and sufficient sample quantity. Quantification of DNAm has been detailed previously [65 (link)]. Research staff extracted genomic DNA from nucleated cells using the PureGene kit (Qiagen, Valencia, CA) and stored sample aliquots at – 80 °C until analysis. DNA was bisulfite converted with the Zymo DNA Methylation kit (Zymo Resarch, Irvine, CA). DNAm was analyzed at Illumina, Inc. with the Illumina 450K Beadchip (Illumina, San Diego, CA), which interrogates > 485,000 methylation sites. To minimize batch effects, 1 μg of DNA from each sample was randomized across plates and BeadChips.
DNAm was measured in all participants with proper consent for genetic and epigenetic analyses and sufficient sample quantity. Quantification of DNAm has been detailed previously [65 (link)]. Research staff extracted genomic DNA from nucleated cells using the PureGene kit (Qiagen, Valencia, CA) and stored sample aliquots at – 80 °C until analysis. DNA was bisulfite converted with the Zymo DNA Methylation kit (Zymo Resarch, Irvine, CA). DNAm was analyzed at Illumina, Inc. with the Illumina 450K Beadchip (Illumina, San Diego, CA), which interrogates > 485,000 methylation sites. To minimize batch effects, 1 μg of DNA from each sample was randomized across plates and BeadChips.
Free full text: Click here
