Expanding the knowledge on dried blood spots and LC-MS-based protein analysis: two different sampling materials and six protein targets.

The combination of dried blood spots (DBS) and bottom-up LC-MS-based protein analysis was investigated in the present paper using six model proteins (1 mg/mL of each protein) with different physicochemical properties. Two different materials for DBS were examined: a water-soluble DBS material (carboxymethyl cellulose, (CMC)) and a commercially available (non-soluble) material (DMPK-C). The sample preparation was optimised regarding the water-soluble material and achieving acceptable repeatability of the signal was emphasised. Five microlitres of whole blood were deposited and dried on either CMC or DMPK-C. The samples were dissolved (CMC) or extracted (DMPK-C) prior to tryptic digest and matrix precipitation. The optimization of the sample preparation showed that an increased buffer concentration (100 mM ammonium bicarbonate) for dissolving the DBS samples gave better repeatability combined with a decrease in analyte signal. CMC seemed to add extra variability (RSD 8-60%) into the analysis compared to sample prepared without CMC (RSD 6-36%), although equal performance compared to DMPK-C material (RSD 13-60%) was demonstrated. The stability of the analytes was examined for different storage periods (1 and 4 weeks) and different storage temperatures (-25, 25, and 40 °C). The stability on both CMC (> 70% compared to reference) and DMPK-C (> 50% compared to reference) was acceptable for most of the peptides. This paper shows that both DBS materials can be used in targeted LC-MS-based protein analysis of proteins with different physicochemical properties. Graphical Abstract Overview of the experimental set-up for expanding the knowledge of dried blood spots in LC-MS-based protein anaysis.