Discover the most talked about and latest scientific content & concepts.

Concept: Protein mass spectrometry


Fast MS techniques have been applied to the analysis of sulfur volatiles in Allium species and varieties to distinguish phenotypes. Headspace sampling by proton transfer reaction (PTR) MS and surface sampling by desorption electrospray ionization (DESI) MS were used to distinguish lachrymatory factor synthase (LFS)-silenced (tearless; LFS-) onions from normal, LFS active (tear-inducing; LFS+), onions. PTR-MS showed lower concentrations of the lachrymatory factor (LF, 3) and dipropyl disulfide 12 from tearless onions. DESI-MS of the tearless onions confirmed the decreased LF 3, and revealed much higher concentrations of the sulfenic acid condensates. Using DESI-MS with MS2 could distinguish zwiebelane ions from thiosulfinate ions. DESI-MS gave reliable fast phenotyping of LFS+ versus LFS- onions by simply scratching leaves and recording the extractable ions for <0.5 min. DESI-MS leaf compound profiles also allowed the rapid distinction of a variety of Allium cultivars to aid plant breeding selections.

Concepts: Mass spectrometry, Electrospray ionization, Taylor cone, Ion source, Electrospray, Protein mass spectrometry, Desorption electrospray ionization, Cultivar


Conventional electrospray ionization mass spectrometry (ESI-MS) uses a capillary for sample loading and ionization. Along with the development of ambient ionization techniques, ESI-MS using noncapillary emitters has attracted more interest in recent years. Following our recent report on ESI-MS using wooden tips (Anal. Chem. 83, 8201-8207 (2011)), the technique was further investigated and extended in this study. Our results revealed that the wooden tips could serve as a chromatographic column for separation of sample components. Sequential and exhaustive ionization was observed for proteins and salts on wooden tips with salts ionized sooner and proteins later. Nonconductive materials that contain microchannels/pores could be used as tips for ESI-MS analysis with sample solutions loaded to the sharp-ends only, since rapid diffusion of sample solutions by capillary action would enable the tips to become conductive. Tips of inert materials such as bamboo, fabrics, and sponge could be used for sample loading and ionization, while samples such as tissue, mushroom, and bone could form tips to induce ionization for direct analysis with application of a high voltage. Figure.

Concepts: Mass spectrometry, Electrospray ionization, Taylor cone, Ion source, Electrospray, Protein mass spectrometry, John Bennett Fenn, Desorption electrospray ionization


In this contribution we present an innovative way to easy, fast and highly sensitive analyses by capillary electrophoresis with electrospray ionization mass spectrometric (ESI-MS) detection. The new method is designed to be applied to ESI-compatible electrolytes (e.g. ammonium acetate) and offers advanced tuning of selectivity conditions within a wide range of analyte mobilities. We use a full capillary isotachophoresis (ITP) format to provide powerful on-line analyte stacking at the ITP boundary all the way to detection and introduce the model of extended ITP where a controlled concentration of the leading ion is added to the terminating zone. Such systems preserve all properties of an ITP system and the velocity of the stacking ITP boundary can be tuned by the composition of both the leading and terminating zone. In this way the system properties can be controlled flexibly and the mobility window of stacked analytes can be tailored to actual needs. The presented theory and the newly defined concept of zone-related boundary mobility allow easy assessment of system selectivity using simple diagrams. We demonstrate the model and its potential on the example of simple acidic cationic systems composed of only two substances (ammonium and acetate) including the example of thiabendazole analysis with a detection limit of 10(-10) M (20 ng/L) and its determination in orange juice by direct sampling after filtration, selective stacking by a tuned extended ITP system and ESI-MS detection.

Concepts: Mass spectrometry, Electrophoresis, Electrospray ionization, Taylor cone, Ion source, Electrospray, Protein mass spectrometry, John Bennett Fenn


Abstract Background: Accumulation of advanced glycation end products (AGEs) in tissues is a major risk factor for diabetes-associated complications. Skin autofluorescence (SAF) values measured by a specific noninvasive approach (AGE Reader; DiagnOptics Technologies B.V., Gröningen, The Netherlands) reflect the overall AGE exposure in skin. Subjects and Methods: In 16 adolescents with type 1 diabetes (age range, 11-18 years) we tested the association between SAF measured with an AGE Reader and the presence of glucuronic acid, 3-indoxyl sulfate, 3-hydroxybutyrate, phenol sulfate, and pentosidine in skin tissue determined with desorption electrospray ionization mass spectrometry (DESI-MS). These compounds are implicated in long-term diabetes complications. Results: SAF values significantly correlated with levels of compounds measured by DESI-MS (r>0.9 and P<0.001 for each). Conclusions: The strong correlation between adolescents' SAF values measured with the AGE Reader and some glycation products measured with DESI-MS indicates that SAF values may be used as surrogate markers of skin exposure to glycemic end products in type 1 diabetes.

Concepts: Mass spectrometry, Diabetes mellitus type 1, Electrospray ionization, Taylor cone, Ion source, Electrospray, Protein mass spectrometry, Desorption electrospray ionization


Drug analysis is an indispensable task in controlling drug abuse, which is a serious problem worldwide nowadays. In this study, we report a simple and rapid approach for detection and quantitation of drugs-of-abuse in urine and oral fluid by wooden-tip electrospray ionization mass spectrometry (WT-ESI-MS). We demonstrated that ketamine, one of the most common abused drugs, and its major metabolite, norketamine, in raw urine and oral fluid could be readily detected and quantified by WT-ESI-MS with only little sample preparation and no chromatographic separation, and the analytical performances, including the linear range, accuracy, precision, LOD and LOQ, were well acceptable for analysis of real samples.

Concepts: Mass spectrometry, Analytical chemistry, Drug addiction, Electrospray ionization, Taylor cone, Ion source, Electrospray, Protein mass spectrometry


A novel test sample for the spatially resolved quantification of illicit drugs on the surface of a fingerprint using time-of-flight secondary ion mass spectrometry (ToF-SIMS) and desorption electrospray ionization mass spectrometry (DESI-MS) was demonstrated. Calibration curves relating the signal intensity to the amount of drug deposited on the surface was generated from inkjet-printed arrays of cocaine, methamphetamine, and heroin with a deposited-mass ranging nominally from 10 pg to 50 ng per spot. These curves were used to construct concentration maps that visualized the spatial distribution of the drugs on top of a fingerprint, as well as being able to quantify the amount of drugs in a given area within the map. For the drugs on the fingerprint on silicon, ToF-SIMS showed great success as it was able to generate concentration maps of all three drugs. On the fingerprint on paper, only the concentration map of cocaine could be constructed using ToF-SIMS and DESI-MS as the signals of methamphetamine and heroin were completely suppressed by matrix and substrate effects. Spatially resolved quantification of illicit drugs using imaging mass spectrometry is possible, but the choice of substrates could significantly affect the results.

Concepts: Mass spectrometry, Drug addiction, Heroin, Electrospray ionization, Taylor cone, Ion source, Protein mass spectrometry, Desorption electrospray ionization


Electrospray ionization mass spectrometry (ESI-MS) is nowadays one of the cornerstones of biomolecular mass spectrometry and proteomics. Advances in sample preparation and mass analyzers have enabled researchers to extract much more information from biological samples than just the molecular weight. In particular, relevant for structural biology, noncovalent protein-protein and protein-ligand complexes can now also be analyzed by MS. For these types of analyses, assemblies need to be retained in their native quaternary state in the gas phase. This initial small niche of biomolecular mass spectrometry, nowadays often referred to as “native MS,” has come to maturation over the last two decades, with dozens of laboratories using it to study mostly protein assemblies, but also DNA and RNA-protein assemblies, with the goal to define structure-function relationships. In this perspective, we describe the origins of and (re)define the term native MS, portraying in detail what we meant by “native MS,” when the term was coined and also describing what it does (according to us) not entail. Additionally, we describe a few examples highlighting what native MS is, showing its successes to date while illustrating the wide scope this technology has in solving complex biological questions. Graphical Abstract ᅟ.

Concepts: Molecular biology, Mass spectrometry, Molecule, Electrospray ionization, Taylor cone, Ion source, Protein mass spectrometry, John Bennett Fenn


Retrospective proteomic studies, including those which aim to elucidate the molecular mechanisms driving cancer, require the assembly and characterization of substantial patient tissue cohorts. The difficulty of maintaining and accessing native tissue archives has prompted the development of methods to access archives of formalin-fixed tissue. Formalin-fixed tissue archives, complete with patient meta data, have accumulated for decades, presenting an invaluable resource for these retrospective studies. This review presents the current knowledge concerning formalin-fixed tissue, with descriptions of the mechanisms of formalin fixation, protein extraction, top-down proteomics, bottom-up proteomics, quantitative proteomics, phospho- and glycoproteomics as well as imaging mass spectrometry. Particular attention has been given to the inclusion of proteomic investigations of archived tumour tissue. This article is part of a Special Issue entitled: Medical Proteomics.

Concepts: Protein, Bioinformatics, Mass spectrometry, Proteomics, Protein mass spectrometry, Top-down proteomics, Shotgun proteomics, Bottom-up proteomics


Mass spectrometry (MS)-based proteomics workflows can crudely be classified into two distinct regimes, either targeting relatively small peptides (i.e. 0.7 kDa < Mw < 3.0 kDa) or small to medium sized intact proteins (i.e. 10 kDa < Mw < 30 kDa), respectively termed bottom-up and top-down proteomics. Recently, a niche has started to be explored covering the analysis of middle-range peptides (i.e. 3.0 kDa < Mw < 10 kDa), aptly termed middle-down proteomics. Although middle-down proteomics can follow, in principle, a modular workflow similiar to that of bottom-up proteomics, we hypothesized that each of these modules would benefit from targeted optimization to improve its overall performance in the analysis of middle-range sized peptides. Hence, to generate middle-range sized peptides from cellular lysates we explored the use of the proteases Asp-N and Glu-C, and a non-enzymatic acid induced cleavage. To increase the depth of the proteome, an SCX separation, carefully tuned to improve the separation of longer peptides, combined with RP-LC using columns packed with material possessing a larger pore size were used. Finally, after evaluating the combination of potentially beneficial MS settings, we also assessed the peptide fragmentation techniques HCD, ETD and EThcD for characterization of middle-range sized peptides. These combined improvements clearly improve the detection and sequence coverage of middle-range peptides and should guide researchers to explore further how middle-down proteomics may lead to an improved proteome coverage, beneficial for, amongst other things, the enhanced analysis of (co-occurring) post-translational modifications.

Concepts: Protein, Bioinformatics, Amino acid, Mass spectrometry, Proteomics, Protein mass spectrometry, Shotgun proteomics, Bottom-up proteomics


The aggregation of the amyloid-β peptide (Aβ) to form fibrils and plaques is strongly associated with Alzheimer’s disease (AD). Although it is well established that this process generates neurotoxicity, it is also heterogeneous with a variety of species being formed during the conversion process. This heterogeneity makes it difficult to detect and characterize each of the aggregates formed, which precludes establishing the specific features responsible for the neurotoxicity observed. Here we use pulse-labeling hydrogen-deuterium exchange experiments analyzed by electrospray ionization mass spectrometry (PL-HDX-ESI-MS) to distinguish three ensembles populated during the aggregation of the 40 and 42 residue forms of the Aβ peptide, Aβ40 and Aβ42, on the basis of differences in their persistent structure. Noticeably, two of them are more abundant at the beginning and at the end of the lag phase and are therefore not detectable by conventional assays such as Thioflavin T (ThT). The ensembles populated at different stages of the aggregation process have a surprisingly consistent average degree of exchange, indicating that there are definite structural transitions between the different stages of aggregation. To determine whether an ensemble of species with a given hydrogen exchange pattern correlates with neurotoxicity, we combined PL-HDX-ESI-MS experiments with parallel measurements of the neurotoxicity of the samples under study. The results of this dual approach show that the maximum toxicity correlates with the ensemble comprising HDX protected oligomers, indicating that development of persistent structure within Aβ oligomers is a determinant of neurotoxicity.

Concepts: Alzheimer's disease, Protein, Mass spectrometry, Structure, Electrospray ionization, Taylor cone, Electrospray, Protein mass spectrometry