Project description:Protein abundance profiling using isobaric labeling is a well-established quantitative mass spectrometry technique. However, ratio distortion resulting from co-isolated and co-fragmented ions - commonly referred to as interference - remains a caveat of this technique. Tribrid mass spectrometers, such as the Orbitrap Fusion and the Orbitrap Fusion Lumos with a triple mass analyzer configuration, facilitate methods (namely SPS-MS3) that can help alleviate interference. Yet, few standards are available to measure interference. Here we introduce the TKO6 standard that assesses ion interference and is designed specifically for data acquired at low (unit) mass resolution. We use TKO6 to compare the degree of interference in MS2 versus MS3-based quantitation methods, data acquisition methods of different lengths, and ion trap-based TMT reporter ion analysis (IT-MS3). We show that the TKO6 standard is a valuable tool for assessing data quality and is particularly useful for benchmarking ion trap-based SPS-MS3 analyses.
Project description:In-solution digests of ankyrin IPs subjected to LC-MS/MS on a Fusion Lumos collecting data in a data-dependent bottoms-up proteomics method.
Project description:Quantitative LC-MS/MS was performed on 3 uL of each sample, using a nanoAcquity UPLC system (Waters Corp) coupled to a Thermo Orbitrap Fusion Lumos high resolution accurate mass tandem mass spectrometer (Thermo) via a nanoelectrospray ionization source. Briefly, the sample was first trapped on a Symmetry C18 20 mm x 180 um trapping column (5 ul/min at 99.9/0.1 v/v water/acetonitrile), after which the analytical separation was performed using a 1.8 um Acquity HSS T3 C18 75 um x 250 mm column (Waters Corp.) with a 60-min linear gradient of 3 to 30% acetonitrile with 0.1% formic acid at a flow rate of 400 nanoliters/minute (nL/min) with a column temperature of 55C. Data collection on the Fusion Lumos mass spectrometer was performed in a data-dependent acquisition (DDA) mode of acquisition with a r=120,000 (m/z 200) full MS scan from m/z 375 - 1500 with a target AGC value of 4e5 ions. MS/MS scans were acquired at Rapid scan rate (Ion Trap) with an AGC target of 1e4 ions and a max injection time of 100 ms. The total cycle time for MS and MS/MS scans was 2 sec. A 20 sec dynamic exclusion was employed to increase depth of coverage. The total analysis cycle time for each sample injection was approximately 1.5 hours.
Project description:Single cell proteomics data containing quantitative information of THP1 and U937 monocytes that were treated or not to undergo a macrophage-like differentiation. Dataset also contains "Mix" samples generated by mixing in equal proportions THP1 and U937 peptides in the single-cell range or by processing together 1 THP1 cell and 1 U937 cell. Samples run on the Orbitrap Fusion Lumos Tribrid and the Exploris 240 were acquired by the de Duve institute, UCLouvain. Samples run on the timsTOF SCP were acquired by the GIGA institute, ULiège.
Project description:In-gel trypsin digested proteins (WNK4 and KLHL3) subjected to LC-MS/MS on a Fusion Lumos collecting data with a ETHcD bottoms-up proteomics method.
Project description:<p>Untargeted features from the Bermuda Atlantic Time-series Study (BATS) site collected during the time period spanning 2016 to 2019. Metabolites were sampled from surface seawater to 1000 m deep and throughout the year. Dissolved organic matter extracts were analyzed in positive and negative ion mode with an ultra-high performance liquid chromatography system (Vanquish UHPLC, Thermo Scientific) coupled with an Orbitrap Fusion Lumos Tribid mass spectrometer.</p>