Conference Report: ‘Large Meets Small‘: Connecting The Bioanalytical Community Around Peptide And Protein Bioanalysis With Lc–Ms(/Ms)

Timmerman, Philip; Lausecker, Berthold; Amsterdam, Peter van; Luedtke, Silke; Dijksman, Jessica

doi:10.4155/bio.12.37

Cited by 12 publications

(6 citation statements)

References 2 publications

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“…Quantitative mass spectrometry (MS) assays for protein biomarkers are typically based on selected or multiple reaction monitoring (SRM or MRM) strategies. Awareness is increasing that such MRM assays need analytical performance criteria which are fit-for-purpose. − In this respect, MS-based protein quantifications for clinical diagnostics demand the highest grade of analytical quality and have been classified as Tier-1 assays or as a field named quantitative Clinical Chemistry Proteomics (qCCP) . In addition, consideration of metrological traceability and the impact of analytical quality on clinical decisions might necessitate even more-stringent criteria for accuracy (defined as trueness and precision) than the guidelines from regulatory agencies .…”

Section: Introductionmentioning

confidence: 99%

Quantifying Protein Measurands by Peptide Measurements: Where Do Errors Arise?

et al. 2015

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Clinically actionable quantification of protein biomarkers by mass spectrometry (MS) requires analytical performance in concordance with quality specifications for diagnostic tests. Laboratory-developed tests should, therefore, be validated in accordance with EN ISO 15189:2012 guidelines for medical laboratories to demonstrate competence and traceability along the entire workflow, including the selected standardization strategy and the phases before, during, and after proteolysis. In this study, bias and imprecision of a previously developed MS method for quantification of serum apolipoproteins A-I (Apo A-I) and B (Apo B) were thoroughly validated according to Clinical and Laboratory Standards Institute (CLSI) guidelines EP15-A2 and EP09-A3, using 100 patient sera and either stable-isotope labeled (SIL) peptides or SIL-Apo A-I as internal standard. The systematic overview of error components assigned sample preparation before the first 4 h of proteolysis as major source (∼85%) of within-sample imprecision without external calibration. No improvement in imprecision was observed with the use of SIL-Apo A-I instead of SIL-peptides. On the contrary, when the use of SIL-Apo A-I was combined with external calibration, imprecision improved significantly (from ∼9% to ∼6%) as a result of the normalization for matrix effects on linearity. A between-sample validation of bias in 100 patient sera further supported the presence of matrix effects on digestion completeness and additionally demonstrated specimen-specific biases associated with modified peptide sequences or alterations in protease activity. In conclusion, the presented overview of bias and imprecision components contributes to a better understanding of the sources of errors in MS-based protein quantification and provides valuable recommendations to assess and control analytical quality in concordance with the requirements for clinical use.

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Section: Introductionmentioning

confidence: 99%

Quantifying Protein Measurands by Peptide Measurements: Where Do Errors Arise?

et al. 2015

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“…For example, currently, a wide range of criteria are being applied in the proteomics community to assert that an assay has been successfully developed and that analytes of interest are being confidently detected and changes in their levels reliably quantified. Proteomics scientists are only slowly implementing practices in targeted, quantitative assay development that have been learned and adopted by the small molecule community (11–13). Equally troubling is that targeted-MS papers are being published without clear and complete documentation of the analytical methods used or the assay performance, making it difficult if not impossible for reviewers and readers to have confidence that the tests work as intended to apply the tests described in their own labs with an expectation that similar results can be achieved.…”

mentioning

confidence: 99%

Targeted Peptide Measurements in Biology and Medicine: Best Practices for Mass Spectrometry-based Assay Development Using a Fit-for-Purpose Approach

Carr

Abbatiello

Ackermann³

et al. 2014

Molecular & Cellular Proteomics

508

546

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Adoption of targeted mass spectrometry (MS) approaches such as multiple reaction monitoring (MRM) to study biological and biomedical questions is well underway in the proteomics community. Successful application depends on the ability to generate reliable assays that uniquely and confidently identify target peptides in a sample. Unfortunately, there is a wide range of criteria being applied to say that an assay has been successfully developed. There is no consensus on what criteria are acceptable and little understanding of the impact of variable criteria on the quality of the results generated. Publications describing targeted MS assays for peptides frequently do not contain sufficient information for readers to establish confidence that the tests work as intended or to be able to apply the tests described in their own labs. Guidance must be developed so that targeted MS assays with established performance can be made widely distributed and applied by many labs worldwide. To begin to address the problems and their solutions, a workshop was held at the National Institutes of Health with representatives from the multiple communities developing and employing targeted MS assays. Participants discussed the analytical goals of their experiments From the ‡Broad Institute of MIT and Harvard, Cambridge, Massachusetts; §Eli

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“…Decades of method optimization established stable isotope dilution as the gold standard for quantitative mass spectrometry . Measurement of the ratio of an analyte to its stable isotope standard (SIS) accounts for fluctuations of analysis and matrix effects .…”

Section: Srm and Mrm Quantificationmentioning

confidence: 99%

Section: Srm and Mrm Quantificationmentioning

confidence: 99%

Targeted methods for quantitative analysis of protein glycosylation

Goldman

Šanda²

2015

Proteomics Clinical Apps

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Quantification of proteins by LC-MS/MS-MRM has become a standard method with broad projected clinical applicability. MRM quantification of protein modifications is, however, far less utilized, especially in the case of glycoproteins. This review summarizes current methods for quantitative analysis of protein glycosylation with a focus on MRM methods. We describe advantages of this quantitative approach, analytical parameters that need to be optimized to achieve reliable measurements, and point out the limitations. Differences between major classes of N- and O-glycopeptides are described and class-specific glycopeptide assays are demonstrated.

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Conference Report: ‘Large Meets Small‘: Connecting The Bioanalytical Community Around Peptide And Protein Bioanalysis With Lc–Ms(/Ms)

Cited by 12 publications

References 2 publications

Quantifying Protein Measurands by Peptide Measurements: Where Do Errors Arise?

Quantifying Protein Measurands by Peptide Measurements: Where Do Errors Arise?

Targeted Peptide Measurements in Biology and Medicine: Best Practices for Mass Spectrometry-based Assay Development Using a Fit-for-Purpose Approach

Targeted methods for quantitative analysis of protein glycosylation

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