Advancement in nanoparticle-based proteomic analysis of whole blood obtained from various dried blood spot collection devices

Microsampling is a methodology for collection and analysis of low-volume biological samples, typically under 100 μL. Dried blood spot (DBS) collected through microsampling affords several advantages compared to conventional venipuncture blood draw: diminished invasiveness, enhanced convenience, and cost-effectiveness. It allows collecting whole blood samples for widespread application in clinical trials, therapeutic drug monitoring, pharmacogenomics and disease diagnosis. While microsampling of DBS has demonstrated significant advantages in proteomics studies, the depth of protein coverage remains limited owing to the high dynamic range of proteins present in whole blood. Also, the biological differences, particularly the differences in hematocrit level, along with low sample volume may cause variations in sample quantification and analysis, which ultimately results in lack of reproducibility of the measurements.

Whole blood samples were obtained intravenously, alongside capillary blood collected via finger-prick from the same subjects to saturate microsampling devices. A simple extraction method was utilized to extract whole blood from microsampling devices, including MitraTM devices (30uL), CapitainerTM devices (10uL), and WhatmanTM paper. The extracted proteins underwent Seer's ProteographTM XT workflow, followed by mass spec analysis using Orbitrap ExplorisTM 480 and AstralTM.

Incubation of the protein extracts from all three devices with the nanoparticles in ProteographTM XT workflow significantly enhanced the number of protein groups detected, compared to the direct processing of the samples. Introducing Seer"s nanoparticles to the extracts from all DBS devices resulted in detection of around 3,000 protein groups on Exploris  480, with additional protein coverage on Astral. In the absence of nanoparticles, the direct processing of the extract from all three microsampling devices resulted in identifying only around 1,000 proteins. The results also exhibited low coefficients of variation (CVs) below 10% across multiple subjects, with similar protein group profiles. The capacity for unbiased proteomic studies at such depth, provided by ProteographTM XT workflow offers a distinct advantage for early biomarker discovery and health status monitoring. Considering the annual collection of millions of dried blood spot (DBS) samples, Seer"s technology can be employed as a powerful tool for accurate and large-scale proteomic studies using DBS samples.