PROTEOMICS

This instrument couples trapped ion mobility spectrometry (IMS) with liquid chromatography (LC) and a time of flight (TOF) mass analyzer. Unique TIMS/PASEF (Parallel Accumulation-Serial Frangmentation) technology allows for the analysis of lower sample amounts, identification and quantitation of near-isobaric peptides and post-translationally modified (PTMs) peptides, including PTM-positional isomers. This results in the transition from 3D-Proteomics (retention time, m/z and MS/MS fingerprint) into 4D-Proteomics (addition of CCS values). 4D-Proteomics enabled by PASEF allows researchers to dig deeper into the proteome and provides more confidence in peptide identification and quantitation in both proteomics and PTM studies by enhancing structural information that is complementary to LC and MS. PASEF enables MS/MS acquisition at >100 Hz, making high-throughput measurements using short gradients. TIMS separates ions on the basis of their shapes and charge state. This complementary mode of mobility separation allows resolution of isomeric peptides (i.e. phosphopeptides and glycopeptides) that differ only by the sites of modification, which provides information about site specific modification dynamics and their relationship to protein structure and functions.

This high-resolution, accurate mass system enables in depth proteome analysis. The instrument can achieve up to 480,000 resolution at m/z 200 and <3 ppm RMS mass accuracy, with a wide mas range, and scan rates up to 40 Hz. A FAIMS Pro interface extends orthogonal sensitivity increases signal-to-noise ratios and expands sample coverage.

Coupled with ETD and U3000 UHPLC system. The Thermo Scientific Orbitrap Fusion Lumos Tribrid Mass Spectrometer is one of the newest, state-of-the-art Orbitrap mass spectrometers that integrates quadrupole, orbitrap and linear ion-trap mass analyzers. This instrument provides multiple dissociation techniques including collision induced (CID), high energy collision induced (HCD), and electron transfer (ETD) dissociations to perform multiple peptide fragmentations in parallel. It provides experimental flexibility for applications such as protein quantitation using isobaric tags, low level post-translational modification (PTM) analysis (e.g. phosphorylation, and glycosylation), and identification of intact proteins, protein complexes, and proteoforms. In addition, this mass spectrometer offers very high resolution, sensitivity and scan speed for rapid acquisition rates for analyzing complex protein samples.

Used with Dionex UltiMate 3000 Nano LC system. The Thermo Scientific Q-Exactive high-field (HF) mass spectrometer combines high-resolution accurate mass (HR/AM) Orbitrap mass analyzer with a high performance quadrupole precursor ion selection to deliver unparalleled speed and sensitivity. This benchtop mass spectrometer is coupled with the Thermo Scientific UltiMate 3000 nano flow liquid chromatography (LC) system for LC-MS/MS analysis. It is highly suitable for a large scale proteomic profiling experiments. The combination of high scan speed, high resolving power and mass accuracy ensures high quality and reproducible data. It is routinely used for high-throughput characterization of proteins from a complex mixture including identification, quantitation, post-translational modification and protein-protein interaction. With the scan speed of up to 18 Hz, dynamic range of 106 and resolving power of up to 240k (FWHM) at m/z 200, this instrument significantly increases the capability to identify thousands of low abundant proteins from a single experiment.

Used with Dionex UltiMate 3000 Nano LC system. The Thermo Scientific TSQ Endura LC-MS is designed for targeted proteomic analysis with highest sensitivity, accuracy, and robustness. This MS comprises three quadrupoles for fast full-scan MS and MS/MS analysis of peptides, and is suitable for ultrafast selected-reaction monitoring (SRM)-assays for absolute protein quantitation. The scan speed of up to 15,000 Da/sec and linear dynamic range of up to 106, allows for sensitive, robust and routine quantitative measurement. This instrument is coupled with the Thermo Scientific UltiMate 3000 nano LC system for LC-MS/MS analysis. SRM assays using this platform allow for the absolute and highly accurate quantitation of hundreds of proteins and biomarkers in a single experiment, and requires very small amount of sample.

Used with Eksigent 425 Nano LC system. The Sciex Triple TOF 5600+ System is the instrument of choice for high-sensitivity and high-resolution applications. With its fast acquisition speeds of up to 10 msec accumulation time, the TripleTOF can handle comprehensive MS/MS quantitation for proteomics applications. The instrument consistently delivers high resolution for high-speed chromatography with up to 100 spectra in a second — even in Information-Dependent Acquisition (IDA) mode. The TripleTOF is a superior instrument that is used for routine accurate mass determination where low abundance analytes are easily discovered, identified and quantified.

This automation platform streamlines the workflow for hydrogen-deuterium exchange (HDX). It offers automation of critical steps including labeling, quenching, digestion, and injection, while maintain a constant temperature to preserve sample integrity.

One of our research scientists can run samples for you. However, if you have a large number of samples or require frequent use of the facility, we can train you to prepare and run your samples. We will also train you in database searches using different search tools, data filtering, and other downstream bioinformatic analyses, so that you can run proteomics experiments with minimum or no supervision.

Yes, students and postdocs who will be working on a project that requires frequent use of an instrument can be trained. To discuss training on an instrument, contact Uma Aryal or Vicki Hedrick.

If you have been trained on an instrument, you can schedule usage in iLab. Any request for services can also be submitted through the iLab system.

Sample requirements vary depending on the type of analysis that you require. Please contact the facility staff to determine how much sample you need for a given analysis or instrument.

Turnaround times can vary. Depending on the number of samples that are submitted and type of analysis required, turnaround usually runs 7 to 10 days.

A Proteomics interest group has been discussed, and there has been some preliminary interest expressed by faculty around campus. If you are interested in such a group, please contact Uma Aryal, director of Purdue Proteomics Facility, at uaryal@purdue.edu or call (765)-494-4960.