Dataset Information

Cov²MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.

ABSTRACT: The pandemic readiness toolbox needs to be extended, targeting different biomolecules, using orthogonal experimental set-ups. Here, we build on our Cov-MS effort using LC-MS, adding SISCAPA technology to enrich proteotypic peptides of the SARS-CoV-2 nucleocapsid (N) protein from trypsin-digested patient samples. The Cov²MS assay is compatible with most matrices including nasopharyngeal swabs, saliva, and plasma and has increased sensitivity into the attomole range, a 1000-fold improvement compared to direct detection in a matrix. A strong positive correlation was observed with qPCR detection beyond a quantification cycle of 30-31, the level where no live virus can be cultured. The automatable sample preparation and reduced LC dependency allow analysis of up to 500 samples per day per instrument. Importantly, peptide enrichment allows detection of the N protein in pooled samples without sensitivity loss. Easily multiplexed, we detect variants and propose targets for Influenza A and B detection. Thus, the Cov²MS assay can be adapted to test for many different pathogens in pooled samples, providing longitudinal epidemiological monitoring of large numbers of pathogens within a population as an early warning system.

SUBMITTER: Van Puyvelde B

PROVIDER: S-EPMC9773173 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature

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Publications

Cov<sup>2</sup>MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.

Van Puyvelde Bart B Van Uytfanghe Katleen K Van Oudenhove Laurence L Gabriels Ralf R Van Royen Tessa T Matthys Arne A Razavi Morteza M Yip Richard R Pearson Terry T Drouin Nicolas N Claereboudt Jan J Foley Dominic D Wardle Robert R Wyndham Kevin K Hankemeier Thomas T Jones Donald D Saelens Xavier X Martens Geert G Stove Christophe P CP Deforce Dieter D Martens Lennart L Vissers Johannes P C JPC Anderson N Leigh NL Dhaenens Maarten M

Analytical chemistry 20221209 50

The pandemic readiness toolbox needs to be extended, targeting different biomolecules, using orthogonal experimental set-ups. Here, we build on our Cov-MS effort using LC-MS, adding SISCAPA technology to enrich proteotypic peptides of the SARS-CoV-2 nucleocapsid (N) protein from trypsin-digested patient samples. The Cov<sup>2</sup>MS assay is compatible with most matrices including nasopharyngeal swabs, saliva, and plasma and has increased sensitivity into the attomole range, a 1000-fold improve ...[more]

PMID: 36490367

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Project description:BackgroundThe non-nucleoside reverse transcriptase inhibitor dapivirine has been evaluated as a topical microbicidal agent to prevent HIV-1 acquisition. Several clinical trials have evaluated the pharmacokinetics of dapivirine when administered as a 25-mg intravaginal ring. Recent studies have focused on the distribution of dapivirine into breast milk. Drug distribution during lactation and breastfeeding can have implications in terms of infant drug exposure. Thus, sensitive bioanalytical tools are required to characterize the pharmacokinetics of dapivirine in breast milk.MethodsWhole breast milk was spiked with dapivirine and internal standard. Lipid content was disrupted via pre-treatment with n-hexane, and supernatants were subjected to solid phase extraction. Extracted materials were subjected to liquid chromatographic-tandem mass spectrometric (LC-MS/MS) analysis. Separation occurred using a Waters BEH C8, 50 × 2.1 mm UPLC column with a 1.7 µm particle size and dapivirine was detected on an API 5000 mass analyzer. Methods were validated in accordance with FDA Bioanalytical Method Validation recommendations.ResultsThe analytical method was optimized for dapivirine extraction from breast milk. The analytical measuring range of the assay was 10-1000 pg/mL. Calibration curves were generated via weighted linear regression of standards. Intra- and inter-assay precision and accuracy studies demonstrated %CVs ≤ 14.6% and %DEVs ≤ ±12.7%. Stability and matrix effects studies were also conducted and deemed acceptable. The method was applied to a previously reported phase 1 clinical trial and demonstrated appropriate performance in the quantitation of dapivirine in breast milk samples from lactating women.ConclusionsAn ultrasensitive LC-MS/MS assay has been developed and validated for the quantitation of dapivirine in breast milk. The described method meets validation acceptance criteria and has been applied to a phase 1 clinical trial.

Dataset Information

Cov²MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.

Publications

Cov<sup>2</sup>MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.

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OmicsDI is part of the ELIXIR infrastructure

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Dataset Information

Cov2MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.

Publications

Cov&lt;sup&gt;2&lt;/sup&gt;MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets

Cov²MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.

Cov<sup>2</sup>MS: An Automated and Quantitative Matrix-Independent Assay for Mass Spectrometric Measurement of SARS-CoV-2 Nucleocapsid Protein.