Fig. 1

Experimental setup. a PBMCs were enriched by Ficoll-assisted centrifugation, exposed to 60 Gy γ-irradiation, and cultured in GMP medium for 24 h. Secretome was filtered, virus-inactivated, and lyophilized (GMP PBMC^sec). To produce placebo control, GMP medium alone without cells was treated equally. b Major biomolecular components present in PBMC^sec (EVs, proteins, and lipids) were analyzed. c A set of physico-chemical-biological tests reflecting key effects mediated by PBMC^sec was selected to determine the reproducibility of 3 PBMC secretome batches produced under GMP. In parallel, tests were performed with one placebo batch. d Selected proteins were quantified and functional assays were performed to compare variability of individual donor secretomes, small, and large secretome batches. e Stability of 2 PBMC^sec batches was assessed after storage at various ambient temperatures for up to 6 months. EVs, extracellular vesicles; GMP, good manufacturing practice; P, phosphorylation; PBMC^sec, secretome of γ-irradiated, peripheral blood mononuclear cells