Analyzing Dried Blood Spots
Dried blood spots (DBS) preserved on filter paper represent a complex matrix used in drug development and newborn blood screening. DBS are an easy way to collect, transport, and store blood samples for testing without the need for refrigeration.
Analyzing compounds in the blood sample can be challenging due to matrix effects caused by the complex makeup of blood and by the components of the paper. Joseph Stankovich, an intern in Gary Van Berkel’s lab at Oak Ridge National Laboratory, described an automated, high-throughput technique called liquid extraction surface analysis (LESA) using Advion BioSystems’ TriVersa Nanomate® nano-electrospray ionization (nano-ESI) MS system for analyzing drugs in DBS.
The success of MS analysis following direct liquid sample extraction from the DBS will depend on the solubility of the analyte and the ionization efficiency, as well as the characteristics of the surface, including the size, shape, topography, and hydrophobicity of the blood spot.
Several techniques have been used to facilitate liquid extraction of DBS, including coating the surface of the filter paper with silicone, creating wells on the surface (for example, by printing a wax pattern on the paper, followed by heating, causing the wax to form shallow wells), or punching out intact spots and putting the filter paper disks directly in the wells of a microtiter plate where the extraction then takes place.
Stankovich showed that all three of these methods yielded comparable results in terms of analyte quantification (in this instance of the drug propranolol), down to 10 ng/mL with an internal standard spiked directly into the liquid blood before spotting. Propranolol was quantified down to a level of 20 ng/mL with addition of an internal standard in methanol on top of the spots after the blood had dried—simulating a practical laboratory situation.
The third technique, punching out the DBS, was the only one that allowed the researchers to apply an internal standard after the initial application of dosed blood and to achieve acceptable accuracy and reproducibility.
Stankovich and colleagues concluded that LESA is a viable method for automated analysis of DBS, and that several techniques can be used to facilitate sample extraction. LESA enabled quantitative sample analysis in less than two minutes per sample.