Concentration Determination of >200 Proteins in Dried Blood Spots for Biomarker Discovery and Validation.

Autor:	Eshghi A; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada. Electronic address: azad@proteincentre.com., Pistawka AJ; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada., Liu J; Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada., Chen M; Island Medical Program, Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, British Columbia V6T 2B5, Canada., Sinclair NJT; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada., Hardie DB; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada., Elliott M; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada., Chen L; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada., Newman R; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada., Mohammed Y; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada; Center for Proteomics and Metabolomics, Leiden University Medical Center, Albinusdreef 2, 2333 ZA Leiden, The Netherlands., Borchers CH; University of Victoria - Genome BC Proteomics Centre, University of Victoria, Victoria, British Columbia V8Z 7X8, Canada; Department of Biochemistry and Microbiology, University of Victoria, Victoria, British Columbia V8P 5C2, Canada; Segal Cancer Proteomics Centre, Lady Davis Institute, Jewish General Hospital, McGill University, Montreal, Quebec H3T 1E2, Canada; Gerald Bronfman Department of Oncology, Jewish General Hospital, McGill University, Montreal, Quebec H3T 1E2, Canada; Department of Data Intensive Science and Engineering, Skolkovo Institute of Science and Technology, Skolkovo Innovation Center, Nobel St., Moscow143026, Russia. Electronic address: christoph.borchers@mcgill.ca.
Jazyk:	angličtina
Zdroj:	Molecular & cellular proteomics : MCP [Mol Cell Proteomics] 2020 Mar; Vol. 19 (3), pp. 540-553. Date of Electronic Publication: 2020 Jan 02.
DOI:	10.1074/mcp.TIR119.001820
Abstrakt:	The use of protein biomarkers as surrogates for clinical endpoints requires extensive multilevel validation including development of robust and sensitive assays for precise measurement of protein concentration. Multiple reaction monitoring (MRM) is a well-established mass-spectrometric method that can be used for reproducible protein-concentration measurements in biological specimens collected via microsampling. The dried blood spot (DBS) microsampling technique can be performed non-invasively without the expertise of a phlebotomist, and can enhance analyte stability which facilitate the application of this technique in retrospective studies while providing lower storage and shipping costs, because cold-chain logistics can be eliminated. Thus, precise, sensitive, and multiplexed methods for measuring protein concentrations in DBSs can be used for de novo biomarker discovery and for biomarker quantification or verification experiments. To achieve this goal, MRM assays were developed for multiplexed concentration measurement of proteins in DBSs.The lower limit of quantification (LLOQ) was found to have a median total coefficient of variation (CV) of 18% for 245 proteins, whereas the median LLOQ was 5 fmol of peptide injected on column, and the median inter-day CV over 4 days for measuring endogenous protein concentration was 8%. The majority (88%) of the assays displayed parallelism, whereas the peptide standards remained stable throughout the assay workflow and after exposure to multiple freeze-thaw cycles. For 190 proteins, the measured protein concentrations remained stable in DBS stored at ambient laboratory temperature for up to 2 months. Finally, the developed assays were used to measure the concentration ranges for 200 proteins in twenty same sex, same race and age matched individuals. (© 2020 Eshghi et al.)
Databáze:	MEDLINE
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