Total and isoform‐specific quantitative assessment of circulating fibulin‐1 using selected reaction monitoring MS and time‐resolved immunofluorometry

Overgaard, Martin; Cangemi, Claudia; Jensen, Martin L.; Argraves, W. Scott; Rasmussen, Lars Melholt

doi:10.1002/prca.201400070

Cited by 7 publications

(12 citation statements)

References 38 publications

(51 reference statements)

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“…Taken together, we conclude that the MRM assay used in this study is fit for purpose but that further development of more focused marker panels with internal digestion controls would be valuable. A clear limitation of the method per se is its relatively low analytical sensitivity for serum or plasma (low μg/ml) when used without any prior reduction of sample complexity [26,36]. Nevertheless, targeted MS offers an interesting analytical platform for future multimarker-based prenatal screening.…”

Section: Discussionmentioning

confidence: 99%

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First-trimester multimarker prediction of gestational diabetes mellitus using targeted mass spectrometry

et al. 2016

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Aims/hypothesis Gestational diabetes mellitus (GDM) is associated with an increased risk of pre-eclampsia, macrosomia and the future development of type 2 diabetes mellitus in both mother and child. Although an early and accurate prediction of GDM is needed to allow intervention and improve perinatal outcome, no single protein biomarker has yet proven useful for this purpose. In the present study, we hypothesised that multimarker panels of serum proteins can improve firsttrimester prediction of GDM among obese and non-obese women compared with single markers. Methods A nested case-control study was performed on firsttrimester serum samples from 199 GDM cases and 208 controls, each divided into an obese group (BMI ≥27 kg/m 2 ) and a non-obese group (BMI <27 kg/m 2 ). Based on their biological relevance to GDM or type 2 diabetes mellitus or on their previously reported potential as biomarkers for these diseases, a number of proteins were selected for targeted nano-flow liquid chromatography (LC) MS analysis. This resulted in the development and validation of a 25-plex multiple reaction monitoring (MRM) MS assay. Results After false discovery rate correction, six proteins remained significantly different (p<0.05) between obese GDM patients (n=135) and BMI-matched controls (n=139). These included adiponectin, apolipoprotein M and apolipoprotein D. Multimarker models combining protein levels and clinical data were then constructed and evaluated by receiver operating characteristic (ROC) analysis. For the obese, nonobese and all GDM groups, these models achieved marginally higher AUCs compared with adiponectin alone. Conclusions/interpretation Multimarker models combining protein markers and clinical data have the potential to predict women at a high risk of developing GDM.

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Section: Discussionmentioning

confidence: 99%

“…Sample preparation and MRM-MS analysis Samples were prepared for MRM analysis essentially as described by Overgaard et al (for a detailed description, see ESM Methods) [26]. Briefly, serum samples were diluted 1:20 in 50 mmol/l ammonium bicarbonate and 15 μl samples were denatured, reduced, alkylated and trypsinised.…”

Section: Methodsmentioning

confidence: 99%

First-trimester multimarker prediction of gestational diabetes mellitus using targeted mass spectrometry

et al. 2016

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“…Several studies have been carried out to identify and quantify allele expression in human body fluids using SRM assay. Overgaard et al quantified cardiovascular biomarker fibulin-1 and its circulating isoforms in human plasma [14]. They used bioinformatics analysis to predict total and isoform-specific tryptic peptides, and quantified the absolute amount of total fibulin-1, isoform-1C, and -1D using SRM assay combined with stable isotope dilution.…”

Section: Detection Of Protein Isoforms For Diagnosis and Prognosis Ofmentioning

confidence: 99%

Impact of allele‐specific peptides in proteome quantification

Snyder

2015

Proteomics Clinical Apps

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Mass spectrometry-based proteome technologies have greatly improved our ability to detect and quantify proteomes across various biological samples. High throughput bottom-up proteome profiling in combination with targeted mass spectrometry method, e.g. selected reaction monitoring (SRM) assay, is emerging as a powerful approach in the field of biomarker discovery. In the past few years, increasing number of studies have attempted to integrate genomic and proteomic data for biomarker discovery. Here we describe how allele-specific peptide can be applied in biomarker discovery and their impact in protein quantification.

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“…8–10 In addition, SRM can quantify site-specific protein isoforms, protein truncation, and post-translational modifications (PTMs) from which antibodies often cannot distinguish with high specificity. 9,11,12 However, SRM still suffers from insufficient sensitivity for precise quantification of low-abundance proteins in complex biological samples such as human blood plasma/serum or tissues. Conventional LC-SRM has a good linearity for protein concentrations at the range of ~4–5 orders of magnitude but only allows quantification of proteins at micrograms per milliliter levels in plasma/serum without immunoaffinity depletion of high-abundance proteins or other front-end sample processing.…”

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confidence: 99%

“…3 However, this method shares typical shortcomings of the antibody-based methods (e.g., relatively low multiplexing, unavailability of antibodies for many proteins, difficulties in generating high-quality antibodies for protein modifications, or isoforms). 3,9 Recently, we developed an antibody-independent targeted MS method PRISM-SRM (high-pressure, high-resolution separations with intelligent selection and multiplexing SRM) which utilizes effective chromatographic enrichment for highly sensitive quantification of target proteins in complex samples. 15,20 PRISM-SRM enables quantification of plasma proteins at the 50–100 pg/mL levels when combined with IgY14 depletion, or at sub-nanogram per milliliter to low nanograms to milliliter levels without immunoaffinity depletion.…”

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confidence: 99%

Deep-Dive Targeted Quantification for Ultrasensitive Analysis of Proteins in Nondepleted Human Blood Plasma/Serum and Tissues

et al. 2017

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Mass spectrometry-based targeted proteomics (e.g., selected reaction monitoring, SRM) is emerging as an attractive alternative to immunoassays for protein quantification. Recently we have made significant progress in SRM sensitivity for enabling quantification of low nanograms per milliliter to sub-naograms per milliliter level proteins in nondepleted human blood plasma/serum without affinity enrichment. However, precise quantification of extremely low abundance proteins (e.g., ≤ 100 pg/mL in blood plasma/serum) using targeted proteomics approaches still remains challenging, especially for these samples without available antibodies for enrichment. To address this need, we have developed an antibody-independent deep-dive SRM (DD-SRM) approach that capitalizes on multidimensional high-resolution reversed-phase liquid chromatography (LC) separation for target peptide separation and enrichment combined with precise selection of target peptide fractions of interest, significantly improving SRM sensitivity by ~5 orders of magnitude when compared to conventional LC-SRM. Application of DD-SRM to human serum and tissue provides precise quantification of endogenous proteins at the ~10 pg/mL level in nondepleted serum and at <10 copies per cell level in tissue. Thus, DD-SRM holds great promise for precisely measuring extremely low abundance proteins or protein modifications, especially when high-quality antibodies are not available.

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Total and isoform‐specific quantitative assessment of circulating fibulin‐1 using selected reaction monitoring MS and time‐resolved immunofluorometry

Cited by 7 publications

References 38 publications

First-trimester multimarker prediction of gestational diabetes mellitus using targeted mass spectrometry

First-trimester multimarker prediction of gestational diabetes mellitus using targeted mass spectrometry

Impact of allele‐specific peptides in proteome quantification

Deep-Dive Targeted Quantification for Ultrasensitive Analysis of Proteins in Nondepleted Human Blood Plasma/Serum and Tissues

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