Exploiting ion-mobility mass spectrometry for unraveling proteome complexity.

Ion mobility spectrometry-mass spectrometry (IMS-MS) is experiencing rapid growth in proteomic studies, driven by its enhancements in dynamic range and throughput, increasing the quantitation precision, and the depth of proteome coverage. The core principle of ion mobility spectrometry is to separate ions in an inert gas under the influence of an electric field based on differences in drift time. This minireview provides an introduction to IMS operation modes and a description of advantages and limitations is presented. Moreover, the principles of trapped IMS-MS (TIMS-MS), including parallel accumulation-serial fragmentation are discussed. Finally, emerging applications linked to TIMS focusing on sample throughput (in clinical proteomics) and sensitivity (single-cell proteomics) are reviewed, and the possibilities of intact protein analysis are discussed.

The clinical potential of prm-PASEF mass spectrometry.

INTRODUCTION: The continuous technical improvement in sensitivity and specificity placed mass spectrometry as an alternative method for analyzing clinical samples. In parallel to the rapid development of discovery proteomics, targeted acquisition has been implemented as a complementary option for measuring a small set of proteins with high sensitivity and robustness in a large sample cohort. The combination of trapped ion mobility with a rapid time-of-flight (TOF) mass spectrometer improves the sensitivity even further and triggers the development of prm-PASEF. AREAS COVERED: This article discusses the development of prm-PASEF and its advantages over the existing targeted and discovery methods for analyzing clinical samples. We are also highlighting the different requirements for the use of prm-PASEF on clinical samples. EXPERT OPINION: prm-PASEF takes advantage of a dual ion-mobility trap enabling highly multiplexed targeted acquisition. It allows the implementation of a short chromatographic separation setup without sacrificing the number of targeted peptides. Analyzing clinical samples by prm-PASEF holds the promise to significantly improve throughput while maintaining sensitivity to detect the selected target proteins.

Mass-Spectrometric Detection of SARS-CoV-2 Virus in Scrapings of the Epithelium of the Nasopharynx of Infected Patients via Nucleocapsid N Protein.

The detection of viral RNA by polymerase chain reaction (PCR) is currently the main diagnostic tool for COVID-19 ( Eurosurveillance 2019, 25 (3), 1). The PCR-based test, however, shows limited sensitivity, especially in the early and late stages of disease development ( Nature 2020, 581, 465-469; J. Formosan Med. Assoc. 2020, 119 (6) 1123), and is relatively time-consuming. Fast and reliable complementary methods for detecting the viral infection would be of help in the current pandemic conditions. Mass spectrometry is one of such possibilities. We have developed a mass-spectrometry-based method for the detection of the SARS CoV-2 virus in nasopharynx epithelial swabs based on the detection of the viral nucleocapsid N protein. Our approach shows confident identification of the N protein in patient samples, even those with the lowest viral loads, and a much simpler preparation procedure. Our main protocol consists of virus inactivation by heating and the addition of isopropanol and tryptic digestion of the proteins sedimented from the swabs followed by MS analysis. A set of unique peptides, produced as a result of proteolysis of the nucleocapsid phosphoprotein of SARS-CoV-2, is detected. The obtained results can further be used to create fast parallel mass-spectrometric approaches for the detection of the virus in the nasopharyngeal mucosa, saliva, sputum and other physiological fluids.

Metabolomics reveals the potential metabolic mechanism of infliximab against DSS-induced acute and chronic ulcerative colitis.

Inflammatory bowel disease (IBD) is often accompanied by metabolic imbalance, and infliximab (IFX) can alleviate IBD symptoms, but its metabolic mechanisms remain unclear. To investigate the relationship between IBD, metabolism, and IFX, an acute and chronic ulcerative colitis (UC) model induced by dextran sulfate sodium (DSS) was established. Plasma samples were analyzed using ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry, followed by multivariate statistical analysis. The results showed that IFX could alleviate colonic shortening and reduce colonic pathological damage in acute and chronic mouse colitis, improve acute and chronic UC, and ameliorate metabolic disturbances. Among the 104 elevated metabolites and 170 decreased metabolites, these metabolites mainly belonged to amino acids, glucose, and purines. The changes in these metabolites were mainly associated with drug metabolism-other enzymes, riboflavin metabolism, phenylalanine, tyrosine and tryptophan biosynthesis, phosphonate and phosphinate metabolism, and phenylalanine metabolism. In summary, this study provides a valuable approach to explore the metabolic mechanisms of IFX in treating acute and chronic UC from a metabolomics perspective.

Linkage-specific identification and quantification of sialylated glycans by TIMS-TOF MS through conjugation with metal complexes.

Mass spectrometry is an indispensable technology for the characterization of glycans. However, specific identification of isomeric glycans especially sialylated glycan isomers using mass spectrometry alone is challenging, which is why orthogonal techniques are needed. Aiming to achieve simple, rapid, and specific identification of sialyl-linkage isomers, we reported herein a trapped ion mobility spectrometry time of flight mass spectrometry (TIMS-TOF MS) method for linkage-specific identification of sialylated glycans through conjugation with metal complexes. Two pairs of sialyl-linkage isomers including 3'/6'-sialyllactose (3'/6'-SL) and 3'/6'-sialyl-N-acetyllactosamine (3'/6'-SLN) conjugated with the diethylenetriamine (DETA) or 2,2'; 6',2″-terpyridine (Terpy) ligand and transition metal ion (Mn2+, Fe2+, Co2+, Ni2+, Cu2+, or Zn2+) were studied by TIMS-TOF MS. The two pairs of sialylated isomers were successfully separated with a metal-ligand system, and relative quantification of sialyl-linkage isomers was demonstrated. In addition, the linkage of the sialic acid moiety can also be distinguished with MS/MS in combination with the metal-ligand system.

Disturbed Plasma Lipidomic Profiles in Females with Diffuse Large B-Cell Lymphoma: A Pilot Study.

Lipidome dysregulation is a hallmark of cancer and inflammation. The global plasma lipidome and sub-lipidome of inflammatory pathways have not been reported in diffuse large B-cell lymphoma (DLBCL). In a pilot study of plasma lipid variation in female DLBCL patients and BMI-matched disease-free controls, we performed targeted lipidomics using LC-MRM to quantify lipid mediators of inflammation and immunity, and those known or hypothesised to be involved in cancer progression: sphingolipids, resolvin D1, arachidonic acid (AA)-derived oxylipins, such as hydroxyeicosatetraenoic acids (HETEs) and dihydroxyeicosatrienoic acids, along with their membrane structural precursors. We report on the role of the eicosanoids in the separation of DLBCL from controls, along with lysophosphatidylinositol LPI 20:4, implying notable changes in lipid metabolic and/or signalling pathways, particularly pertaining to AA lipoxygenase pathway and glycerophospholipid remodelling in the cell membrane. We suggest here the set of S1P, SM 36:1, SM 34:1 and PI 34:1 as DLBCL lipid signatures which could serve as a basis for the prospective validation in larger DLBCL cohorts. Additionally, untargeted lipidomics indicates a substantial change in the overall lipid metabolism in DLBCL. The plasma lipid profiling of DLBCL patients helps to better understand the specific lipid dysregulations and pathways in this cancer.