Effect of Anticoagulant and Storage Conditions on Bovine Nonesterified Fatty Acid and β-Hydroxybutyrate Concentrations in Blood

The objective of this study was to determine if nonesterified fatty acid (NEFA) and β-hydroxybutyrate (BHBA) concentrations were affected by anticoagulants or gel and clot activator tubes (serum separator tubes, SST), storage of samples as whole blood, separated plasma or serum at 24°C or 4°C for 24 to 72 h, or storage as serum at −40°C for 1 mo. Blood was collected from dairy cows into EDTA, heparin, nonanticoagulant tubes, and SST, and analyzed immediately to obtain baseline NEFA and BHBA concentrations. Portions were stored as whole blood or separated plasma or serum at 4 or 24°C and assayed daily for 24 (whole blood) and 72 (separated samples) h. Serum samples were frozen at −40°C and assayed at 24 h or weekly for 1 mo. Baseline NEFA concentrations were unaffected by anticoagulants; however, they were significantly higher in SST compared with nonanticoagulant tubes. Concentrations of NEFA were stable in all samples at 4°C, whereas they sequentially increased from 24 to 48 h at 24°C. Changes were more dramatic in heparinized samples. Serum could be stored frozen for up to 1 mo with minimal changes in NEFA concentrations. Concentrations of BHBA were stable under all conditions evaluated. Our results indicated that blood for NEFA testing should be collected into EDTA or nonanticoagulant tubes (but not SST), separated promptly from cells, and maintained at 4°C until analysis.