All-in-one paper-based sampling chip for targeted protein analysis

Skjærvø, Øystein; Halvorsen, Trine Grønhaug; Reubsaet, Léon

doi:10.1016/j.aca.2019.08.043

Cited by 18 publications

(23 citation statements)

References 88 publications

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“…In addition to these significant advances enabling the in-depth characterization of the human plasma and serum proteome, other technological developments have focused on improving the reproducibility, throughput, and sample preparation prior to LC-MS/MS analyses. To circumvent lengthy, error-prone, and variable manual handling, sophisticated chips or microfluidic devices integrating biochemical steps of preanalytical protocols have been developed. − Some of these microfluidic devices integrate plasma preparation, depletion of abundant proteins, digestion, and peptide desalting for a fully automated processing of blood samples . Others developed highly parallelized preanalytical workflows based on automation using the 96-well plate format. − In line with such progress and to avoid bias in the distribution of samples before sample processing, dedicated applications have been developed to randomize samples, supporting the reliability of the entire workflow.…”

Section: Recent Advancesmentioning

confidence: 99%

Advances and Utility of the Human Plasma Proteome

et al. 2021

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The study of proteins circulating in blood offers tremendous opportunities to diagnose, stratify, or possibly prevent diseases. With recent technological advances and the urgent need to understand the effects of COVID-19, the proteomic analysis of blood-derived serum and plasma has become even more important for studying human biology and pathophysiology. Here we provide views and perspectives about technological developments and possible clinical applications that use mass-spectrometry(MS)- or affinity-based methods. We discuss examples where plasma proteomics contributed valuable insights into SARS-CoV-2 infections, aging, and hemostasis and the opportunities offered by combining proteomics with genetic data. As a contribution to the Human Proteome Organization (HUPO) Human Plasma Proteome Project (HPPP), we present the Human Plasma PeptideAtlas build 2021-07 that comprises 4395 canonical and 1482 additional nonredundant human proteins detected in 240 MS-based experiments. In addition, we report the new Human Extracellular Vesicle PeptideAtlas 2021-06, which comprises five studies and 2757 canonical proteins detected in extracellular vesicles circulating in blood, of which 74% (2047) are in common with the plasma PeptideAtlas. Our overview summarizes the recent advances, impactful applications, and ongoing challenges for translating plasma proteomics into utility for precision medicine.

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Section: Recent Advancesmentioning

confidence: 99%

Advances and Utility of the Human Plasma Proteome

et al. 2021

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“…The recent evolution of the MS instrumentation has also enabled high sensitivity and low detection limits and the technique is now able to quantify femtograms of proteins from e.g. cell- 53 or exosome extracts 54 and picograms of proteins in serum and whole blood samples 55 . MS has also vastly contributed to the ongoing positive results and WB failed to identify recombinant SIRPA in the calibrators.…”

Section: Mass Spectrometry In Protein Analysismentioning

confidence: 99%

Instant on-paper protein digestion during blood spot sampling

Skjærvø

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Halvorsen

et al. 2017

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A concept integrating sampling and protein digestion is introduced here combining fast and simple fabrication by wax printing on filter paper with trypsin immobilized polymer beads. The paper reactors showed promising results with a high degree of protein digestion within fifty minutes in model protein mixtures as well as in human blood. The model protein mixture was used for the evaluation of performance both with and without a reduction and alkylation step. The paper reactors without reduction and alkylation showed between 46% and 75% protein sequence coverage and between five and 20 high confidence peptides (one and five zero missed cleavage peptides, respectively). Compared to a conventional in-solution approach, the paper reactor showed 10% less protein sequence coverage, 29% fewer high confidence peptides and 19% fewer high confidence peptides with zero missed cleavages. Placement of the protein reduction and alkylation step (before or after protein digestion) was shown to be of low importance. The storage stability of the paper reactors with (six weeks) and without (twelve weeks) tryptic peptides was satisfactory. The ability of the paper reactors to digest complex biological samples was investigated by comparison with human whole blood samples prepared using a conventional dried blood spot (DBS) procedure with overnight digestion in non-targeted analysis. The reactors showed a comparable performance with 75 ± 25 for the protein groups compared to 76 ± 5 for the DBS samples. Additionally, 267 ± 72 and 335 ± 11 unique peptides (high confidence) were identified for on-paper digestion and DBS, respectively.

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“…Functionalization using HEMA-VDM: For the HEMA-VDM functionalization, the procedure published by Skjaervø et al [29,[36][37][38] was slightly modified and used. In short, the fibers on the paper discs were first silanized by applying 10 µL of freshly prepared silanizing solution (this is a mixture of 2.5 mg DPPH, 330 mg DMF and 172 mg γ-MAPS prepared in a glass vial).…”

Section: Functionalization Of Paper Discs and Protein Immobilizationmentioning

confidence: 99%

“…The long-term goal of our research is to simplify MS-based protein analysis from dried matrix samples (such as DBS), and we aim to integrate time-consuming procedures within the sampling device for a prompt start to the sample preparation upon sample collection [29,[35][36][37][38].…”

Section: Introductionmentioning

confidence: 99%

Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper

et al. 2021

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This paper describes smart sampling paper to be used for bottom-up protein analysis. Four different manners to immobilize trypsin on cellulose were evaluated. Untreated paper, potassium-periodate-functionalized paper (with and without post-immobilization reduction) and 2-hydroxyethyl methacrylate (HEMA)/2-vinyl-4,4-dimethylazlactone (VDM)-functionalized paper were all used to immobilize trypsin. For the evaluation, Coomassie Brilliant Blue staining of proteins on paper and the BAEE trypsin activity assay needed to be modified. These methods allowed, together with data from mass spectrometric analysis of cytochrome C digestions, us to acquire fundamental insight into protein binding, and trypsin action and activity on paper. All functionalized discs bind more protein than the untreated discs. Protein binding to functionalized discs is based on both adsorption and covalent binding. Trypsin immobilized on potassium-periodate-functionalized discs exhibits the highest trypsin activity when using cytochrome C as substrate. It is proven that it is trypsin attached to paper (and not desorbed trypsin) which is responsible for the enzyme activity. The use of discs on complex biological samples shows that all functionalized discs are able to digest diluted serum; for the best-performing disc, HEMA-VDM functionalized, up to 200 high-confidence proteins are qualified, showing its potential.

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All-in-one paper-based sampling chip for targeted protein analysis

Cited by 18 publications

References 88 publications

Advances and Utility of the Human Plasma Proteome

Advances and Utility of the Human Plasma Proteome

Instant on-paper protein digestion during blood spot sampling

Next-Generation Dried Blood Spot Samplers for Protein Analysis: Describing Trypsin-Modified Smart Sampling Paper

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