The pre-analytical stability of 25-hydroxyvitamin D: storage and mixing effects

Background: There is an increasing demand for serum 25-OH VitD testing globally and this has led to the greater use of automated immunoassays. These may be more prone to nonspecific interference, that is thought to be related to pre-analytical stability of biological samples. We have investigated the changes in serum 25-OH VitD concentrations that are caused by storage and mixing conditions, and if such changes are statistical, or clinically important. Methods: Blood samples were collected into plain tubes from 31 healthy donors. After separation, serum samples were stored at -20C° and analysis was carried out with and without mixing (vortexing) at different time intervals of days (0,1,2,3,4,5,15, and 30). All samples were analyzed using a chemiluminescent immunoassay. Results: Mean serum 25-OH VitD concentrations for subsequent days of storage compared to day 0 showed a significant time effect (p< 0.05) except for the samples on day 1 (p=0.654) in non-vortexed samples and day 2 (p=0.087), 5 (p=0.118) and 30 (p=0.118) in vortexed samples. Comparing values for vortexed and non-vortexed samples on the same day, serum 25-OH VitD showed a significant difference on days 1 (p=0.003), 4 (p=0.037), 5 (p=0.002) and 30 (p=0.025). However, the maximum change value was 8.85% which was less than the known total allowable error (TEa) and reference change value (RCV) for serum 25-OH VitD. Conclusion: 25-OH VitD is pre-analytically stable after long-term sample storage at -20°C and can be analyzed without vortexing. This may be beneficial for both research and diagnostic laboratories.