Clinical proteomic biomarker discovery using blood plasma is limited by the presence of many highly abundant proteins. Single-shot LC-MS of undepleted plasma typically yields 200-300 proteins so strategies involving depletion of abundant proteins and peptide fractionation have been necessary to regularly exceed this barrier. These strategies are not recommended for clinical studies that require analysis of hundreds of samples as they introduce variance due to sample processing and consume significant acquisition time, have increased costs and complicate data analysis.
LC-MS of whole blood is not favoured due to the very high concentrations of haemoglobin and structural proteins in red blood cell membranes. Proteomic analyses of dried blood spots have been reported but typically offer no improvement over undepleted plasma analysis.
We explored the use of volumetric absorptive microsampling (VAMS) device (MITRA, Neoteryx Inc.) which can collect microliter volumes of blood from a fingerprick with high repeatability. The VAMS device enables highly abundant blood proteins to be depleted through washing, while retaining other less abundant proteins, including those commonly detected from undepleted plasma. We showed that up to 1,600 proteins can be robustly detected in single-shot DDA runs with acceptable CV% <15% using an Orbitrap HFX with 90min LC gradient. Various washing reagents were tested to establish optimal conditions. We applied the VAMS devices to analyse whole blood pellets which had been frozen for many years, demonstrating utility of this approach for archival specimens which would be otherwise refractory to conventional proteomic analysis. Use of VAMS devices may be of interest for biomarker discovery as it offers the ability to sample plasma proteins and cellular proteins originating from shed somatic cells, leukocytes and erythrocytes in a simple, single-shot workflow from a small volume of self-collected donor blood.