scholarly journals Large-scale human skin lipidomics by quantitative, high-throughput shotgun mass spectrometry

2017 ◽  
Vol 7 (1) ◽  
Author(s):  
Tomasz Sadowski ◽  
Christian Klose ◽  
Mathias J. Gerl ◽  
Anna Wójcik-Maciejewicz ◽  
Ronny Herzog ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Human Skin ◽  
High Throughput ◽  
Large Scale ◽  
Shotgun Mass Spectrometry

scholarly journals High-Throughput Stool Metaproteomics: Method and Application to Human Specimens

mSystems ◽  
2020 ◽  
Vol 5 (3) ◽  
Author(s):  
Carlos G. Gonzalez ◽  
Hannah C. Wastyk ◽  
Madeline Topf ◽  
Christopher D. Gardner ◽  
Justin L. Sonnenburg ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
Clinical Studies ◽  
Large Scale ◽  
Dietary Intervention ◽  
Negative Impact ◽  
Mass Spectrometry Analysis ◽  
Careful Study ◽  
Stool Samples ◽  
Study Participants

ABSTRACT Stool-based proteomics is capable of significantly augmenting our understanding of host-gut microbe interactions. However, compared to competing technologies, such as metagenomics and 16S rRNA sequencing, it is underutilized due to its low throughput and the negative impact sample contaminants can have on highly sensitive mass spectrometry equipment. Here, we present a new stool proteomic processing pipeline that addresses these shortcomings in a highly reproducible and quantitative manner. Using this method, 290 samples from a dietary intervention study were processed in approximately 1.5 weeks, largely done by a single researcher. These data indicated a subtle but distinct monotonic increase in the number of significantly altered proteins between study participants on fiber- or fermented food-enriched diets. Lastly, we were able to classify study participants based on their diet-altered proteomic profiles and demonstrated that classification accuracies of up to 89% could be achieved by increasing the number of subjects considered. Taken together, this study represents the first high-throughput proteomic method for processing stool samples in a technically reproducible manner and has the potential to elevate stool-based proteomics as an essential tool for profiling host-gut microbiome interactions in a clinical setting. IMPORTANCE Widely available technologies based on DNA sequencing have been used to describe the kinds of microbes that might correlate with health and disease. However, mechanistic insights might be best achieved through careful study of the dynamic proteins at the interface between the foods we eat, our microbes, and ourselves. Mass spectrometry-based proteomics has the potential to revolutionize our understanding of this complex system, but its application to clinical studies has been hampered by low-throughput and laborious experimentation pipelines. In response, we developed SHT-Pro, the first high-throughput pipeline designed to rapidly handle large stool sample sets. With it, a single researcher can process over one hundred stool samples per week for mass spectrometry analysis, conservatively approximately 10× to 100× faster than previous methods, depending on whether isobaric labeling is used or not. Since SHT-Pro is fairly simple to implement using commercially available reagents, it should be easily adaptable to large-scale clinical studies.

scholarly journals Obesity Proteomics: An Update on the Strategies and Tools Employed in the Study of Human Obesity

2018 ◽  
Vol 7 (3) ◽  
pp. 27 ◽  
Author(s):  
Afshan Masood ◽  
Hicham Benabdelkamel ◽  
Assim Alfadda
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
Large Scale ◽  
Protein Identification ◽  
Biochemical Characterization ◽  
Biological Fluids ◽  
Protein Composition ◽  
Cellular Protein ◽  
Human Obesity ◽  
Two Dimensional Gel Electrophoresis

Proteomics has become one of the most important disciplines for characterizing cellular protein composition, building functional linkages between protein molecules, and providing insight into the mechanisms of biological processes in a high-throughput manner. Mass spectrometry-based proteomic advances have made it possible to study human diseases, including obesity, through the identification and biochemical characterization of alterations in proteins that are associated with it and its comorbidities. A sizeable number of proteomic studies have used the combination of large-scale separation techniques, such as high-resolution two-dimensional gel electrophoresis or liquid chromatography in combination with mass spectrometry, for high-throughput protein identification. These studies have applied proteomics to comprehensive biochemical profiling and comparison studies while using different tissues and biological fluids from patients to demonstrate the physiological or pathological adaptations within their proteomes. Further investigations into these proteome-wide alterations will enable us to not only understand the disease pathophysiology, but also to determine signature proteins that can serve as biomarkers for obesity and related diseases. This review examines the different proteomic techniques used to study human obesity and discusses its successful applications along with its technical limitations.

scholarly journals Identification of an Arginase II Inhibitor via RapidFire Mass Spectrometry Combined with Hydrophilic Interaction Chromatography

2018 ◽  
Vol 24 (4) ◽  
pp. 457-465 ◽  
Author(s):  
Wataru Asano ◽  
Yu Takahashi ◽  
Motoaki Kawano ◽  
Yoshiji Hantani
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
High Throughput Screening ◽  
Large Scale ◽  
Hydrophilic Interaction Chromatography ◽  
Label Free ◽  
Hydrophilic Interaction ◽  
High Concentration ◽  
Allosteric Site ◽  
Arginase Ii

Peripheral arterial disease (PAD) is an occlusive disease that can lead to atherosclerosis. The involvement of arginase II (Arg II) in PAD progression has been proposed. However, no promising drugs targeting Arg II have been developed to date for the treatment of PAD. In this study, we established a method for detecting the activity of Arg II via high-throughput label-free RapidFire mass spectrometry using hydrophilic interaction chromatography, which enables the direct measurement of l-ornithine produced by Arg II. This approach facilitated a robust high-concentration screening of fragment compounds and the identification of a fragment that inhibits the activity of Arg II. We further confirmed binding of the fragment to the potential allosteric site of Arg II using a surface plasmon resonance assay. We concluded that the identified fragment is a promising compound that may lead to novel drugs to treat PAD, and our method for detecting the activity of Arg II can be applied to large-scale high-throughput screening to identify other structural types of Arg II inhibitors.

scholarly journals Automated high-throughput proteome and phosphoproteome analysis using paramagnetic bead technology

2019 ◽  
Author(s):  
Mario Leutert ◽  
Ricard A. Rodriguez-Mias ◽  
Noelle K. Fukuda ◽  
Judit Villén
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
Large Scale ◽  
Magnetic Particles ◽  
Network Inference ◽  
Mapk Pathway ◽  
Mitogen Activated Protein Kinase ◽  
Kinase Substrate ◽  
Data Independent Acquisition ◽  
Automated Method

AbstractRecent developments in proteomics have enabled signaling studies where >10,000 phosphosites can be routinely identified and quantified. Yet, current analyses are limited in throughput, reproducibility, and robustness, hampering experiments that involve multiple perturbations, such as those needed to map kinase-substrate relationships, capture pathway crosstalks, and network inference analysis. To address these challenges, we introduce rapid-robotic-phosphoproteomics (R2-P2), an end-to-end automated method that uses magnetic particles to process protein extracts to deliver mass spectrometry-ready phosphopeptides. R2-P2 is robust, versatile, high-throughput, and achieves higher sensitivity than classical protocols. To showcase the method, we applied it, in combination with data-independent acquisition mass spectrometry, to study signaling dynamics in the mitogen-activated protein kinase (MAPK) pathway in yeast. Our results reveal broad and specific signaling events along the mating, the high-osmolarity glycerol, and the invasive growth branches of the MAPK pathway, with robust phosphorylation of downstream regulatory proteins and transcription factors. Our method facilitates large-scale signaling studies involving hundreds of perturbations opening the door to systems-level studies aiming to capture signaling complexity.

scholarly journals Large-Scale Structural Characterization of Drug and Drug-Like Compounds by High-Throughput Ion Mobility-Mass Spectrometry

2017 ◽  
Vol 89 (17) ◽  
pp. 9023-9030 ◽  
Author(s):  
Kelly M. Hines ◽  
Dylan H. Ross ◽  
Kimberly L. Davidson ◽  
Matthew F. Bush ◽  
Libin Xu
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
Structural Characterization ◽  
Ion Mobility ◽  
Large Scale ◽  
Ion Mobility Mass Spectrometry

Acoustic Ejection Mass Spectrometry: A Fully Automatable Technology for High-Throughput Screening in Drug Discovery

2021 ◽  
pp. 247255522110281
Author(s):  
Roman P. Simon ◽  
Tim T. Häbe ◽  
Robert Ries ◽  
Martin Winter ◽  
Yuting Wang ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Drug Discovery ◽  
High Throughput ◽  
High Throughput Screening ◽  
Large Scale ◽  
Method Development ◽  
Accurate Determination ◽  
Adenosine Monophosphate ◽  
Guanosine Monophosphate ◽  
Label Free

Acoustic droplet ejection (ADE)–open port interface (OPI)–mass spectrometry (MS) has recently been introduced as a versatile analytical method that combines fast and contactless acoustic sampling with sensitive and accurate electrospray ionization (ESI)–MS-based analyte detection. The potential of the technology to provide label-free measurements in subsecond analytical cycle times makes it an attractive option for high-throughput screening (HTS). Here, we report the first implementation of ADE-OPI-MS in a fully automated HTS environment, based on the example of a biochemical assay aiming at the identification of small-molecule inhibitors of the cyclic guanosine monophosphate–adenosine monophosphate (GMP-AMP) synthase (cGAS). First, we describe the optimization of the method to enable sensitive and accurate determination of enzyme activity and inhibition in miniaturized 1536-well microtiter plate format. Then we show both results from a validation single-concentration screen using a test set of 5500 compounds, and the subsequent concentration–response testing of selected hits in direct comparison with a previously established matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) readout. Finally, we present the development of an in-line OPI cleaning procedure aiming to match the instrument robustness required for large-scale HTS campaigns. Overall, this work points to critical method development parameters and provides guidance for the establishment of integrated ADE-OPI-MS as HTS-compatible technology for early drug discovery.

Rapid Identification of Commercial Frankincense Products by MALDI-TOF Mass Spectrometry

2021 ◽  
Vol 24 ◽  
Author(s):  
Shang-Chih Lai ◽  
Ren-In You ◽  
Tz-Ting Chen ◽  
Yu Chang ◽  
Chao-Zong Liu ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
High Throughput ◽  
High Throughput Screening ◽  
Large Scale ◽  
Reference Sample ◽  
Maldi Tof Ms ◽  
Boswellic Acids ◽  
Maldi Tof ◽  
Tof Ms

Background: Frankincense is a resin secreted by the Boswellia tree. It is used in perfumery, aromatherapy, skincare, and traditional Chinese medicine. However, all Boswellia species are under threat owing to habitat loss and overexploitation. As a result, the market is getting flooded with counterfeit frankincense products. Objective: This study aims to establish a high-throughput method to screen and identify the authenticity of commercial frankincense products. We report, for the first time, a matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based method for rapid and high-throughput screening of frankincense samples. Methods: MALDI-TOF MS, HPLC, thin-layer chromatography (TLC), and in vitro anti-inflammatory activity assay were used to examine the frankincense samples. Results: Well-resolved peaks of frankincense triterpenoids in the spectra were observed in the crude extract of commercial samples, including α-boswellic acids (αBAs), β-boswellic acids (βBAs), 11-keto-β-boswellic acids (KBAs), acetyl-11-keto-β-boswellic acids (AKBAs), and their esters. These compounds can be used as indicators for determining the authenticity of frankincense. Conclusion: Unlike LC–MS, which is a time-consuming and expensive method, and TLC, which requires a reference sample, our inexpensive, rapid high-throughput identification method based on MALDI-TOF MS is ideal for large-scale screening of frankincense samples sold in the market.

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