Is the stability of folates in dried blood microsamples sufficient to perform home-sampling studies?

Dried blood microsampling is increasingly used for home-sampling and epidemiological studies because of its multiple advantages, including an often greatly improved analyte stability. However, a critical assessment of the stability under realistic conditions should always be performed as part of the validation, especially for unstable molecules like folates (vitamin B9). Here, the objective was to determine whether folate stability in dried blood microsamples is sufficient to allow the set-up of home-sampling studies for the monitoring of folate status in e.g., women of reproductive age. An extensive set of stability experiments was performed to evaluate the stability of the main folate vitamer 5-methyltetrahydrofolate (5MTHF), its oxidation product MeFOX and the minor non-methyl folate vitamers 10-formylfolic acid (10FoFA), 5,10-methenyltetrahydrofolate (5,10CH+THF) and tetrahydrofolate (THF) in dried blood microsamples using volumetric absorptive microsampling (VAMS) or regular dried blood spots (DBS). The evaluations included (EDTA-anticoagulated blood was collected from a single donor measured in four replicates per condition and time point): (i) the effect of temperature (-20 °C, 4 °C, ambient temperature and 37 °C), (ii) the effect of light (during drying and storage) and humidity, and (iii) the effect of storage under vacuum and pretreatment of the microsamples with stabilizing agents on folate stability. At -20 °C and 4 °C, all folate levels were within 85 to 115% of the baseline value up till two weeks of storage in both VAMS samples and DBS. However, at room temperature the stability of the analyzed folates was only consistently observed up till three days in VAMS samples, and for none of the folates at 37 °C. Humidity had a major impact on 5,10CH+THF stability, but this could be easily improved by using desiccant. Both vacuum treatment and pretreatment of microsamples with 0.1% DL-dithiothreitol and 5% butylated hydroxytoluene improved the stability at room temperature in VAMS samples, but these effects were limited at 37 °C and in DBS. Overall, the stability of the individual folate vitamers proved to be challenging and strongly temperature- and time-dependent. Nonetheless, if controlled transport (temperature and duration) can be assured, the set-up of home-sampling studies to evaluate the folate status using dried blood microsamples can still be beneficial.