A B C D E F G RNA analysis, 4 plaques Protein analysis, 5 plaques Supplemental Figure 1. Manual grinding of atherosclerotic plaque preserved in RNAlater® into fine powder (GEMS). Sample was maintained at all times submerged in RNAlater® solution until processed. Before grinding a ceramic mortar and pestle were pre-chilled on dry-ice (A) and a sample was allowed to freeze in the vessel (B). Sample was first carefully crushed into smaller pieces (C) before grinding into a finer powder (D). For RNA isolation the powder was overlaid with TRIzol® solution (E). For protein isolation the powder was transferred into a tube containing extraction buffer (F). If required the frozen powder can be preserved for later analysis. For a detailed protocol see Materials and Methods section. (G) A general scheme of the experimental design for evaluation of compositional equivalence of sample aliquots. Intact Tissue Manual pulverization of tissue Isolation of RNA followed by analysis and comparison between aliquots ~10 mm Tissue extraction A B C D E F RNA extraction Protein and lipid extraction G Divide powder into aliquots Isolation of protein followed by analysis and comparison between aliquots Isolation of lipids followed by analysis and comparison between aliquots RNA analysis, 4 plaques Protein analysis, 5 plaques Lipid analysis, 10 plaques Plaque powder see Figure 3A see Figure 3B see Figure 3C