Genechip mogene 1.0 st microarrays

CD8⁺ T cells were enriched with magnetic beads and CD45.2 OT1 CD8⁺ T cells were sorted on a FACSAria (BD Biosciences). RNA was isolated using TRIzol (Invitrogen) according to the manufacturer's instructions. RNA was processed, amplified, labeled, and hybridized to Affymetrix GeneChip MoGene 1.0 ST microarrays (Santa Clara, CA) at the Molecular Profiling Facility of the University of Pennsylvania. Human CMV-specific (HLA-A*0201-NLVPMVATV or HLA-B*0702-TPRVTGGGAM tetramer-positive) CD8⁺ T cells were purified from PBMC obtained from subjects with persistent HCV viremia (infected for > 1year) or from healthy volunteers who were recruited at Massachusetts General Hospital in Boston (Table S1). The study was approved by the local IRB (Protocol # 1999-P-004983/54; MGH #: 90-7246). RNA was isolated, processed, amplified, labeled, and hybridized to Affymetrix Human Gene 1.0 ST microarrays. Affymetrix Power Tools software (Santa Clara, CA) was used to process and quantile normalize fluorescent hybridization signals using the Robust Multichip Averaging method (Irizarry et al., 2003 (link)). Transcripts were log₂ normalized. Hierarchical Clustering was performed with Gene Pattern (Reich et al., 2006 (link)) and Gene Set Enrichment Analysis was performed with GSEA software (Subramanian et al., 2005 (link)).