Dataset Information

Integrating serum protein electrophoresis with mass spectrometry, a new workflow for M-protein detection and quantification

ABSTRACT: Serum protein electrophoresis (SPE) and immunofixation electrophoresis (IFE) are standard tools for multiple myeloma (MM) routine diagnostics. M-protein is a biomarker for MM that can be quantified with SPE and characterized with IFE. We have investigated combining SPE/IFE with targeted mass spectrometry (MS) to detect and quantify the M-protein. SPE-MS assay offers the possibility to detect M-protein with higher sensitivity than SPE/IFE, which could lead to better analysis of minimal residual disease in clinical laboratories. In addition, analysis of archived SPE gels could be used for retrospective MM studies. We have investigated two different approaches of measuring M-protein and therapeutic monoclonal antibodies (t-mAbs) from SPE/IFE gels. After extracting proteotypic peptides from the gel, they can be quantified using stable isotope labeled (SIL) peptides and measured by Orbitrap mass spectrometry. Alternatively, extracted peptides can be labeled with tandem mass tags (TMT). Both approaches are not hampered by the presence of t-mAbs. Using SIL peptides, limit of detection of the M-protein is approximately 100-fold better than with routine SPE/IFE. Using TMT labeling, M-protein can be measured and compared in different samples from the same patient. We have successfully measured M-protein proteotypic peptides extracted from the SPE/IFE gels utilizing SIL peptides and TMT.

INSTRUMENT(S): Orbitrap Fusion, Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood

SUBMITTER: Marina Zajec

LAB HEAD: Theo Luider

PROVIDER: PXD016325 | Pride | 2020-01-07

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	180323HFes_JS-1664-dara-A_033.dat	Other
	180323HFes_JS-1664-dara-A_033.raw	Raw
	180323HFes_JS-1664-dara-B_032.dat	Other
	180323HFes_JS-1664-dara-B_032.raw	Raw
	180323HFes_JS-1664-dara-C_031.dat	Other

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Publications

Integrating Serum Protein Electrophoresis with Mass Spectrometry, A New Workflow for M-Protein Detection and Quantification.

Zajec Marina M Jacobs Joannes F M JFM de Kat Angelino Corrie M CM Dekker Lennard J M LJM Stingl Christoph C Luider Theo M TM De Rijke Yolanda B YB VanDuijn Martijn M MM

Journal of proteome research 20200114 7

Serum protein electrophoresis (SPE) and immunofixation electrophoresis (IFE) are standard tools for multiple myeloma (MM) routine diagnostics. M-protein is a biomarker for MM that can be quantified with SPE and characterized with IFE. We have investigated combining SPE/IFE with targeted mass spectrometry (MS) to detect and quantify the M-protein. SPE-MS assay offers the possibility to detect M-protein with higher sensitivity than SPE/IFE, which could lead to better analysis of minimal residual d ...[more]

PMID: 31895568

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Project description:Targeted mass spectrometry (MS) approaches, which are powerful methods for uniquely and confidently quantifying a specific panel of proteins in complex biological samples, play a crucial role in validation and clinical translation of protein biomarkers discovered by global proteomic profiling. Common targeted MS methods, such as multiple reaction monitoring (MRM) and parallel-reaction monitoring (PRM), employ specific mass spectrometric technologies to quantify protein levels by comparing the transitions of specific endogenous (Endo) peptides with those of stable isotope labeled (SIL) peptide counterparts. These methods utilizing amino acid analyzed (AAA) SIL peptides warrants sensitive and precise measurements required for targeted MS assays. Compared to MRM, PRM provides higher experimental throughput by simultaneously acquiring all transitions of the target peptides and thereby compensate for different ion suppression among transitions of a target peptide. However, PRM still suffers different ion suppressions between Endo and SIL peptides due to poor spray stability as the Endo and SIL peptides were monitored at different liquid chromatography (LC) retention times. Here we introduce a new targeted MS method, termed as wideband PRM (WBPRM), designed for high-throughput targeted MS approach. WBPRM employs a wide isolation window for simultaneous fragmentations of both Endo and SIL peptides along with multiplexed single ion monitoring (SIM) scans for enhanced MS sensitivity of target peptides. Compared to PRM, WBPRM was demonstrated to provide increased sensitivity, precision, and accuracy of quantitative measurements of target peptides with increased throughput, allowing more target peptide measurements in a short experiment time. Its adaptability to a MS method provided by the manufacture makes it a facile method to implement for MS based assays, particularly in complex biological samples, where the needs for higher accuracy, sensitivity and efficiency are paramount.