Case Study: Measuring Biomarkers in Dried Blood Spot Samples
There has been a recent upsurge in interest for using dried blood spot (DBS) samples for the clinical measurement of biomarkers. DBS technology was implemented clinically in the 1970s for neonatal screening and has since been expanded to many other applications.
Benefits of DBS sampling include a small blood volume requirement, easy sample collection, no centrifugation steps, and simplified storage and transport. However, the sensitivity of DBS tests is often poor, and most assay formats that use DBS samples are tedious and have limited throughput. Sensitivity and sample-size issues have, until now, limited the utility of DBS sampling for measuring low-abundance biomarkers.
NanoInk has combined the benefits of a 2 µL sample size requirement with a DBS elution protocol to develop a miniaturized immunoassay capable of measuring multiple biomarkers in small-volume samples with improved sensitivity. A multiplexed cytokine DBS assay has been fully validated and shown to have low well-to-well variability, low slide-to-slide variability, and excellent standard and QC recoveries.
For this validation study, the NanoArrayer 3000 was used to print cytokine capture antibodies in 48 different sub-arrays of a functionalized glass slide. The resulting feature sizes were between 2.5–3 µm in diameter.
DBS standards and QC samples were eluted in an elution buffer prior to analysis in the multiplex NanoArray assay (Figure 2) using streptavidin-Alexa Fluor® 647 as a detection molecule. DBS accuracy and precision validation statistics were determined for three different runs over two days. The multiplexed cytokine DBS assay passed all validation criteria.
In addition, NanoInk has conducted preliminary correlation studies with DBS and serum samples from rheumatoid arthritis patients for a selection of biomarkers (including TNF-α, leptin, MIP-1b, IP-10, IL-8, and IFNγ). The DBS and serum samples collected thus far have been tested in a nanoarray multiplex assay, and the concentrations of biomarkers were determined. This study is still under way, but preliminary results suggest a significant DBS-serum correlation for a panel of biomarkers.
This case study has confirmed that DBS sampling can provide significant benefits over conventional blood draws. The nanoarray assay has been validated using DBS samples in a multiplex format, and we have demonstrated that the NanoInk assay can provide a significant amount of biomarker data with minimum sample volumes.