Long-Term Stability of 18 Nutritional Biomarkers Stored at −20 °C and 5 °C for up to 12 Months

Background: Consistent information on long-term storage stability for a broad range of nutritional biomarkers is lacking. We investigated the stability of 18 biomarkers stored at suboptimal temperatures (−20 °C and 5 °C) for up to 12 months. Methods: Multiple vials of serum or whole blood pools (3 concentrations) were stored at −20 °C or 5 °C, removed from the −20 °C freezer after 3, 6, 9, and 12 months and from the 5 °C refrigerator after 6 and 12 months, and placed into a −70 °C freezer until analysis at study completion. Vials stored continuously at −70 °C were used as the reference condition for optimal storage. We measured 18 biomarkers: 4 iron status, 1 inflammation, 8 water-soluble vitamin, and 5 fat-soluble vitamin. For each temperature, we calculated geometric mean concentrations and average percent changes of geometric means across pools relative to the reference condition estimated from a linear mixed model. Results: Most biomarkers (13 of 18) showed no difference in concentration after 12 months of storage at −20 °C. Serum ferritin (1.5%), soluble transferrin receptor (−1.7%), and folate (−10.5%) showed small to moderate significant changes at 6 months, but changes were acceptable based on biologic variability. Serum pyridoxal-5′-phosphate (−18.6% at 9 months) and vitamin C (−23% at 6 months) showed large and unacceptable changes at −20 °C. All serum fat-soluble vitamins and iron status indicators, vitamin B12, total homocysteine, and methylmalonic acid showed acceptable changes when stored at 5 °C for up to 12 months. Conclusions: Overall, we found good long-term stability for multiple nutritional biomarkers stored at suboptimal temperatures.