Total RNA was extracted from frozen brain samples using the Ambion RNAqueous kit according to the manufacturer's instructions. The quantity and quality of the RNA samples were determined by the Agilent 2100 Bioanalyzer using the Agilent RNA 6000 Nano Chip. Transcript levels were measured using the Whole Genome DASL assay (Illumina, San Diego, CA). Probe annotations were done based on NCBI Ref Seq, Build 36.2. The RNA samples were randomized across the chips and plates using a stratified approach to ensure balance with respect to diagnosis, age, gender, RINs and APOE genotype. Replicate samples were utilized for QC and also for intra-class coefficient (ICC) estimations. Raw probe level mRNA expression data were exported from GenomeStudio software (Illumina Inc.) for preprocessing with background correction, variance stabilizing transformation, quantile normalization and probe filtering using the lumi package of BioConductor [56], [57] (Text S1). A probe with detectable signal in >75% of the samples was regarded as informative and used in subsequent analyses, although we also did supplementary analyses without imposing any restrictions based on probe detection levels. The number of informative probes differed slightly between the AD, non–AD and combined groups (Figure S7).
