MASS SPECTROMETRY CORE FACILITY
, and , Co-Directors
Nancy E. Vieira, MS, Senior Research Assistant
The NICHD Mass Spectrometry Facility was created under the auspices of the Office of the Scientific Director to provide high-end mass-spectrometric services to all NICHD scientists, particularly in the area of protein characterization. The facility is located in the 9th D corridor of Building 10 of NIH’s Bethesda campus.
Mode of Operation
Facility staff are available for consultation to everyone within the Institute free of charge, with the provision that existing resources are distributed equally among investigators requesting our services. With the exception of the single-analyzer TOF instrument, no instrument is available for general use. The philosophy of the Mass Spectrometry Facility is to ensure that only reliable, high-quality data are recorded on its instruments and that only statistically meaningful analyses are provided to its clients. For every new project, staff members meet with the principal investigator and postdoctoral scientists involved in the study to discuss details of the experimental design. We discuss the background of the project and analytical goals to determine the most appropriate techniques for sample preparation before bringing the samples to the laboratory. Upon delivery of protein samples, typically in the form of gel separated by the investigator, proteins are digested in situ, and the resulting peptides are isolated for mass-spectrometric analysis. We subject separated peptides to analysis in which selected peptides are fragmented by a second stage of mass spectrometer operation. We then use the fragmentation spectra as input to database search engines to determine the protein(s) present in the original sample. Upon completion of an analysis, we discuss results with the investigators. Long-term user feedback indicates that high-level interaction greatly improves the efficiency of each project.
Instrumentation
The facility currently has four mass spectrometers used for specific areas of analysis.
(1) ABI 4800 MALDI TOF/TOF. This state-of-the-art high-performance instrument can be operated in both positive and negative ion modes. We typically use it for peptide mixtures without chromatographic separation. It is also used for accurate determination of peptide masses in complex mixtures and for the determination of sequences of unusual peptides through techniques of de novo sequencing.
(2) Thermo Finnigan LCQ Deca and LCQ DecaXP. These instruments employ liquid chromatography and electrospray ionization to separate and ionize peptides that are then fragmented.
(3) ABI Voyager MALDI TOF. This instrument is suitable for analysis of higher–molecular weight species—equal to or less than 125 kDa—and for verification of the molecular weight of intact proteins. It is also available for general use, after a potential user has been appropriately trained.
Community Outreach
The Mass Spectrometry Facility is committed to promoting mass-spectrometric aspects of proteomics in the NICHD, Division of Intramural Research. We educate investigators on the benefits and pitfalls of the techniques used in the facility by (1) coaching on the principles of appropriate methods for isolating and staining gels and (2) supporting an NIH-wide seminar series that features nationally known experts in proteomics. In parallel to these efforts, the staff have developed collaborations with other Institutes to promote the exchange of information and to bring new mass-spectrometric techniques to the NICHD.
Facility Usage
The Mass Spectrometry Facility currently serves 15 sections within the Institute, representing about 40 projects as well as four principal investigators of sister Institutes on the campus.
For further information, contact aly@helix.nih.org.
