ILSB Mass Spectrometry Lab (ILSB-MSL)

Mass spectrometry plays a key role in molecular level research and is central to ‘omics’ research, i.e., proteomics, metabolomics, lipidomics, glycomics, as well as materials research, nutrition, drug discovery, reproductive biology, environmental chemistry and biology. The ILSB-MSL is staffed by highly skilled technical staff and state-of-the-art instrumentation to provide advanced MS research capabilities to support a broad range of molecular-level research and develop projects and train scientists to advance the field of mass spectrometry to new levels. The ILSB-MSL specializes MS in peptide mass mapping, protein ID, proteomics and lipidomics, as well as advanced research capabilities that challenge routine technologies, viz. ultra-high resolution MS, ion mobility-MS, and native MS/IMMS in support of studies of large proteins and membrane protein complexes. Challenging research problems often require development of new methodologies and/or modifications of existing approaches; consequently, many of these projects are handled as collaborative research projects. We strongly encourage researchers to consult with us about their project first, before samples are submitted for analysis.

Faculty seeking access to mass spectrometry analysis are often confused by the fact that there are other MS laboratories and instruments located in the ILSB, specifically the Laganowsky and Russell research groups; however, these units are not formally part of the CBMSC. In addition, the Texas A&M NIH P41 (funded July 1, 2018) “Resource in Native Mass Spectrometry-Guided Structural Biology”, which includes laboratories at Ohio State University (lead unit) and West Virginia University, provide unique, cutting-edge mass spectrometry capabilities. The Laganowsky and Russell research programs serve as “Driving Biological Projects” (Laganowsky) and “Technology Research and Development Programs” (Russell) as components of the P41 grant. The NIH P41 “Resource” has received partial funding from the Department of Chemistry, College of Science and OVPR; consequently, these research capabilities are made available on a collaborative basis to Texas A&M researchers. Because these instruments are not formally part of the CBMSL, we have no mechanism to recover operational, maintenance and analysis costs. We currently have a number of collaborations that have produced publications in high-impact journals, and these are the kinds of products that serve as the basis for competitive federal grants that enhance Texas A&M’s stature and interdisciplinary training/education of students.

Protein Identification

Protein identification is most commonly accomplished by proteolytic digestion followed by MS analysis. Sample are typically submitted as gel bands, pelleted pure proteins, or proteins in solution without detergents or glycerol (preferably in 25mM Ammonium Bicarbonate buffer, pH=7-8). Samples are enzymatically digested, typically with mass spec grade trypsin, and analyzed by either MALDI-TOF-MS/MS or nano-capillary HPLC/MSMS. The resulting MS and MS/MS data can then be queried against a specific database for peptide and protein identification.

Exact Mass Analysis for Proteins

Clients often need to know the exact mass of a purified protein. Samples need to be in Na and K free buffers, in the absence of detergents, chaotrophic agents, and glycerol. We recommend ammonium acetate or ammonium bicarbonate buffers if they are required.

Intact Protein Complex Mass Determination

Masses of purified protein complexes and subunit composition can also be determined in our facility. This is non-routine work and we highly recommend a consultation before submitting your samples.

Proteomic Analysis

Complex mixtures of proteins can also be identified by MS. However, protein ID requires LCMS for analysis. Prefractionation of the proteins, such as 1D SDS-PAGE can be used to increase the numbers of proteins identified by the experiment. Other approaches include separation of digest proteins peptides using high pH LC, collecting fractions and running those fractions by low pH LCMS. Yet another approach is to run long gradients using normal reverse phase chromatography-MS. Set up a consultation and we can decide which is best for you. We also offer semi-quantitative techniques, either label-free or labeling for your proteomic analysis.

Posttranslational Modification Site Determination

Starting with a single highly purified protein, either as a gel band or purified protein, multiple sites of modification, eg. Glycosylation, phosphorylation, acetylation and others, can be determined. This process is non-routine and typically requires multiple digestion enzymes. We recommend you set up a consultation before beginning this work.

The ILSB-MSL instrument inventory currently includes, Thermo Orbitrap Fusion Tribrid MS coupled UHPLC, Waters Synapt G2 equipped with capabilities for surface-induced dissociation (SID), Agilent 6560 ion mobility-Q-ToF extended mass range (20 kDa m/z) MS equipped with UHPLC, Agilent 6545 Q-TOF with extended mass range (30 kDa) equipped with UHPLC, a one-of-a-kind Fourier-transform IMS-Orbitrap MS (Thermo Exactive EMR extend mass range). These advanced MS instrumentation and highly skilled staff in the ILSB-MSL offers Texas A&M scientists access to more advanced and diverse mass spectrometry research capabilities. The research capabilities of the ILSB-MSL are further augmented by the staff and instrumentation in the NIH P41 “Resource in Native MS Guided Structural Biology”, which is also located in the ILSB.