Journal ArticleDOI
Stop and Go extraction tips for matrix-assisted laser desorption/ionization, nanoelectrospray, and lc/ms sample pretreatment in proteomics
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TLDR
A novel procedure in which a very small disk of beads embedded in a Teflon meshwork is placed as a microcolumn into pipet tips, finding that the Stage system is well-suited as a universal sample preparation system for proteomics.Abstract:
Proteomics is critically dependent on optimal sample preparation. Particularly, the interface between protein digestion and mass spectrometric analysis has a large influence on the overall quality and sensitivity of the analysis. We here describe a novel procedure in which a very small disk of beads embedded in a Teflon meshwork is placed as a microcolumn into pipet tips. Termed Stage, for STop And Go Extraction, the procedure has been implemented with commercially available material (C18 Empore Disks (3M, Minneapolis, MN)) as frit and separation material. The disk is introduced in a simple and fast process yielding a convenient and completely reliable procedure for the production of self-packed microcolumns in pipet tips. It is held in place free of obstacles solely by the narrowing tip, ensuring optimized loading and elution of analytes. Five disks are conveniently placed in 1 min, adding 300 μL/...read more
Citations
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Journal ArticleDOI
Universal sample preparation method for proteome analysis
TL;DR: A method is described, filter-aided sample preparation (FASP), which combines the advantages of in-gel and in-solution digestion for mass spectrometry–based proteomics and allows single-run analyses of organelles and an unprecedented depth of proteome coverage.
Journal ArticleDOI
In-gel digestion for mass spectrometric characterization of proteins and proteomes
TL;DR: This protocol is for the in-gel digestion of both silver and Coomassie-stained protein spots or bands and can be followed by MALDI-MS or LC-MS/MS analysis to identify proteins at sensitivities better than a few femtomoles of protein starting material.
Journal ArticleDOI
Accurate Proteome-wide Label-free Quantification by Delayed Normalization and Maximal Peptide Ratio Extraction, Termed MaxLFQ
TL;DR: A new intensity determination and normalization procedure called MaxLFQ is developed that is fully compatible with any peptide or protein separation prior to LC-MS analysis, which accurately detects the mixing ratio over the entire protein expression range, with greater precision for abundant proteins.
Journal ArticleDOI
Protocol for micro-purification, enrichment, pre-fractionation and storage of peptides for proteomics using StageTips
TL;DR: StageTips are ordinary pipette tips containing very small disks made of beads with reversed phase, cation-exchange or anion-ex exchange surfaces embedded in a Teflon mesh that allow flexible combination of disks with different surfaces to obtain multi-functional tips.
Journal ArticleDOI
Tumour exosome integrins determine organotropic metastasis
Ayuko Hoshino,Bruno Costa-Silva,Tang-Long Shen,Gonçalo Rodrigues,Ayako Hashimoto,Milica Tesic Mark,Henrik Molina,Shinji Kohsaka,Angela Di Giannatale,Sophia Ceder,Swarnima Singh,Caitlin Williams,Nadine Soplop,Kunihiro Uryu,Lindsay A. Pharmer,Tari A. King,Linda Bojmar,Alexander E. Davies,Yonathan Ararso,Tuo Zhang,Haiying Zhang,Jonathan M. Hernandez,Joshua Mitchell Weiss,Vanessa D. Dumont-Cole,Kimberly Kramer,Leonard H. Wexler,Aru Narendran,Gary K. Schwartz,John H. Healey,Per Sandström,Knut Jørgen Labori,Elin H. Kure,Paul M. Grandgenett,Michael A. Hollingsworth,Maria de Sousa,Sukwinder Kaur,Maneesh Jain,Kavita Mallya,Surinder K. Batra,William R. Jarnagin,Mary S. Brady,Øystein Fodstad,Volkmar Müller,Klaus Pantel,Andy J. Minn,Mina J. Bissell,Benjamin A. Garcia,Yibin Kang,Yibin Kang,Vinagolu K. Rajasekhar,Cyrus M. Ghajar,Irina Matei,Héctor Peinado,Jacqueline Bromberg,Jacqueline Bromberg,David Lyden +55 more
TL;DR: It is demonstrated that exosomes from mouse and human lung-, liver- and brain-tropic tumour cells fuse preferentially with resident cells at their predicted destination, namely lung fibroblasts and epithelial cells, liver Kupffer cells and brain endothelial cells.
References
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Laser desorption ionization of proteins with molecular masses exceeding 10,000 daltons
Michael Karas,Franz Hillenkamp +1 more
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Journal ArticleDOI
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Johan Gobom,Johan Gobom,Eckhard Nordhoff,Ekaterina Mirgorodskaya,Rolf Ekman,Peter Roepstorff +5 more
TL;DR: It is shown that 2,5-dihydroxybenzoic acid (DHB) acts as an efficient matrix for peptide mapping and both detection sensitivity and sequence coverage are comparable to those obtained with the currently preferred matrix alpha-cyano-4-hydroxycinnamic acid (CHCA).
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