Smart blood spots for whole blood protein analysis

Abstract

A reactor for whole blood sampling integrated with instant protein digestion in a “lab-on-paper” format is introduced here. The sampling reactor was fabricated on commercially available filter paper using 2-hydroxyethyl methacrylate-co-2-vinyl-4,4-dimethyl azlactone (HEMA-VDM) polymerization followed by the immobilization of trypsin. Immobilization conditions were investigated with respect to temperature, enzyme amount and immobilization time. The highest reactor efficiency with respect to protein digestion was obtained with 1.25 mg trypsin per reactor immobilized at room temperature for 3 hours. Commercially available cellulose filter papers and DMPK-C cards were modified and immobilized with trypsin prior to whole blood sampling. Filter paper specifications including thickness (180–220 μm), weight (77–92 g m⁻²) and porosity (11–25 μm) were investigated with respect to performance (digestion efficiency and extraction recovery). From this study, it was found that a medium thickness paper with higher weight and porosity is optimal for reactor efficiency. The reactors were tested and compared with respect to a standard dried blood spot procedure for protein digestion. The most efficient reactors obtained 134 ± 14 and 124 ± 7 high confidence protein groups for freeze thawed and fresh whole blood samples, respectively.