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Transcriptome changes were de\ufb01ned after 1h treatment with 32\u25e6C hypothermia (hsp70 family member change), 10 \u00b5mol\/L ferric citrate that had no discernible effect, and 100 \u00b5g\/mL cycloheximide leading to induction of primary response (immediate early) genes including IL1B and TNF. Same-donor PBMCs prepared conventionally using washes then resuspension in serum-supplemented media demonstrated basal upregulation of stress signalling pathway genes that masked and overlapped differential gene  expression pro\ufb01les after 100 \u00b5g\/L cycloheximide. Plasma-resuspended PBMCs demonstrated minor transcriptome changes after 40 \u00b5mol\/L ferric citrate, whereas consistent and greater magnitude changes were observed for washed\/media- resuspended PBMCs. We conclude that endogenous plasma-maintained PBMCs provide a more robust platform to interrogate acute cellular perturbations trig- gering innate immunity, and that varying susceptibility of PBMCs to preparative stresses is an important component of experimental variability.<\/jats:p>","DOI":"10.15698\/cst2024.11.301","type":"journal-article","created":{"date-parts":[[2024,12,3]],"date-time":"2024-12-03T11:53:36Z","timestamp":1733226816000},"page":"112-124","source":"Crossref","is-referenced-by-count":3,"title":["Endogenous plasma resuspension of peripheral blood mononuclear cells prevents preparative-associated stress that modi\ufb01es polyA-enriched RNA responses to subsequent acute 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