Quantitation of peptides and proteins by matrix-assisted laser desorption/ionization mass spectrometry using18O-labeled internal standards

Mirgorodskaya, O. A.; Kozmin, Yuri; Титов, М. И.; Körner, Roman; Sönksen, Carsten P.; Roepstorff, Peter

doi:10.1002/1097-0231(20000730)14:14<1226::aid-rcm14>3.0.co;2-v

Cited by 335 publications

(13 citation statements)

References 19 publications

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“…Note that tandem MS identifications have been recently reported also in the absence of detectable precursor signals (Panchaud et al 2009), suggesting that isobaric methods may be more sensitive than isotopic ones. The last approach for differential quantification by chemical derivatization is enzymatic labeling , exemplified by 16 O/ 18 O labeling (Mirgorodskaya et al 2000), where the mass tag is introduced in the peptide chain by performing proteolytic digestion in the presence of heavy water.…”

Section: Stable Isotope-based Quantitative Proteomicsmentioning

confidence: 99%

Current challenges in software solutions for mass spectrometry-based quantitative proteomics

et al. 2012

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Mass spectrometry-based proteomics has evolved as a high-throughput research field over the past decade. Significant advances in instrumentation, and the ability to produce huge volumes of data, have emphasized the need for adequate data analysis tools, which are nowadays often considered the main bottleneck for proteomics development. This review highlights important issues that directly impact the effectiveness of proteomic quantitation and educates software developers and end-users on available computational solutions to correct for the occurrence of these factors. Potential sources of errors specific for stable isotope-based methods or label-free approaches are explicitly outlined. The overall aim focuses on a generic proteomic workflow.

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Section: Stable Isotope-based Quantitative Proteomicsmentioning

confidence: 99%

Current challenges in software solutions for mass spectrometry-based quantitative proteomics

et al. 2012

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“…In this study, we assessed the potential effects of triple fusion (TF) reporter gene expression on embryonic stem (ES) cell function. To accomplish this, we used isotopic labeling of peptides with 16 O/ 18 O [21,22] and observed no significant changes by quantitative protein analysis. Likewise, the TF did not adversely affect ES cell viability, proliferation, and differentiation.…”

Section: Introductionmentioning

confidence: 99%

Proteomic analysis of reporter genes for molecular imaging of transplanted embryonic stem cells

Cao

Dutta

et al. 2006

Proteomics

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Study of stem cells may reveal promising treatment for diseases. The fate and function of transplanted stem cells remain poorly defined. Recent studies demonstrate that reporter genes can monitor real-time survival of transplanted stem cells in living subjects. We examined the effects of a novel and versatile triple fusion (TF) reporter gene construction on embryonic stem (ES) cell function by proteomic analysis. Murine ES cells were stably transduced with a self-inactivating lentiviral vector containing fluorescence (firefly luciferase; Fluc), bioluminescence (monomeric red fluorescence protein; mRFP), and positron emission tomography (herpes simplex virus type 1 truncated thymidine kinase; tTK) reporter genes. Fluorescence-activated cell sorting (FACS) analysis isolated stably transduced populations. TF reporter gene effects on cellular function were evaluated by quantitative proteomic profiling of control ES cells versus ES cells stably expressing the TF construct (ES-TF). Overall, no significant changes in protein quantity were observed. TF reporter gene expression had no effect on ES cell viability, proliferation, and differentiation capability. Molecular imaging studies tracked ES-TF cell survival and proliferation in living animals. In summary, this is the first proteomic study, demonstrating the unique potential of reporter gene imaging for tracking ES cell transplantation non-invasively, repetitively, and quantitatively.

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“…The use of MALDI-TOF MS to measure isotopomer peaks after protein labeling has been explored with yeast and Escherichia coli for various applications including peptide identification [21,22], confirmation of peptide sequencing and calibration of MS [23], and for improving identification of protein modifications [24]. Complete labeling of cells in vitro (Stable Isotope Labeling by Amino acids in Cell culture, SILAC) has been described as a quantitative proteomics approach [25]. Also, labeling during analysis has been employed for protein quantification with MALDI-TOF [26].…”

Section: Discussionmentioning

confidence: 99%

A combination of protein profiling and isotopomer analysis using matrix‐assisted laser desorption/ionization‐time of flight mass spectrometry reveals an active metabolism of the extracellular matrix of 3T3‐L1 adipocytes

2004

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Differential gel electrophoresis followed by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry (MS) is a commonly used protein profiling method. However, observed changes can be explained in multiple ways, one of which is by the protein turnover rate. In order to easily and rapidly obtain information on both the identity and turnover of individual proteins, we applied a combination of protein labeling with L-(ring-2,3,4,5,6 2H5) phenylalanine and MALDI-TOF MS. While the spectrum reveals the identity of a protein, mass isotopomer analysis provides information about the rate of protein labeling as a measure of synthesis or turnover. Using this approach on mature 3T3-L1 adipocytes, we were able to discriminate between rapidly and slowly metabolised proteins. In our isolate, proteins of the cytoskeleton appeared to be slowly metabolised, whereas components of the extracellular matrix, in particular collagen type I alpha 1 (COL1A1) and collagen type I alpha 2 (COL1A2) showed rapid accumulation of newly synthesized proteins. Both proteins appeared to be metabolised in the same ratio as they are present in collagen fibers, i.e. 2:1 (COL1A1: COL1A2). In addition, functionally related proteins were also readily labeled. Taken together, we have shown that a combination of stable isotope labeling and protein profiling by gel electrophoresis and MALDI-TOF analysis can simultaneously provide information on the identity and relative metabolic rate of proteins in eukaryotic cells in a simple, nonhazardous and rapid-throughput way.

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Quantitation of peptides and proteins by matrix-assisted laser desorption/ionization mass spectrometry using18O-labeled internal standards

Cited by 335 publications

References 19 publications

Current challenges in software solutions for mass spectrometry-based quantitative proteomics

Current challenges in software solutions for mass spectrometry-based quantitative proteomics

Proteomic analysis of reporter genes for molecular imaging of transplanted embryonic stem cells

A combination of protein profiling and isotopomer analysis using matrix‐assisted laser desorption/ionization‐time of flight mass spectrometry reveals an active metabolism of the extracellular matrix of 3T3‐L1 adipocytes

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