Plasma collection, handling and storage

Blood collection, processing and storage were performed under strict conditions to minimize pre-analytical variability [6, 20]. Fasting EDTA plasma was separated from whole blood within 2–4 hours of venepuncture and immediately stored at -80°C prior to bio-banking. Samples then undergo a single freeze thaw cycle for the purpose of creating aliquots, which minimizes subsequent freeze thaw cycles for specific experiments. EDTA plasma was chosen as anticoagulant since it chelates divalent metals, thereby protecting plasma constituents from oxidation, which is particularly important for lipids. Thereafter, lipid extractions were performed within 15 minutes of freeze thawing and extracts stored at -80°C.