High Levels of HMGB1 in Plasma May Be due to Ex Vivo Cell Necrosis

a. Levels of HMGB1 in plasma samples exposed to different preanalytical conditions. The conditions included analysis: after 1 hour, overnight (o/n) storage at 4°C or room temperature (RT), vortexing for 20-30 seconds, one cycle of freeze/thawing (F/T). Data are presented as median, interquartile range. p-Value refers to Kruskal-Wallis analysis with Graph Prism Software. (C) -indicates the centrifugation step. b. Western blot analysis of HMGB1 in plasma samples obtained after vortexing (vortex), overnight incubation (o/n) and freeze/thaw (F/T) cycle. Necrotic extract from PBMCs and recombinant HMGB1 (100 ng; R&D systems, MN, USA) served as a control.