Facilities: Proteomics
RCMI Proteomics Core Facility is equipped with Mass Spectrometer , Chromatography, Gel Electrophoresis setups and Molecular Biology research support facilities.
LTQ Orbitrap XL tandem Mass Spectrometer
This mass spectrometer consists of two spectrometers: Linear Quadrupole Trap (LTQ) and Orbitrap. LTQ is a low resolution mass spectrometer used for fast analysis, filtration of the molecules with the masses of interest, and fragmentation of molecules in a trap chamber. These options make LTQ unique for MS-n analysis. Fourier Transform Orbitrap Spectrometer has a unique sensitivity of ~2 ppm. Tandem combination of these two traps results in a top level Mass Spectrometry solution.
Mass spectrometer have a few options for sample molecules fragmentation: Collision Induced Dissociation (CID), Pulsed Q Dissociation (PQD), High Energy Collision Dissociation (HCD).
Analyte ionization is made from liquid phase through Electro Spray Ionization (ESI) or Nano Spray Ionization (NSI). NSI is an electro spray ionization system operating at nanoL/min flow rate. This considerably increases ionization efficiency and sensitivity of the system.
For higher efficiency analyte could be preliminarily separated by HPLC systems. For ESI ionization we use Thermo Finnigan HPLC system. NSI works in tandem with Shimadzu nano HPLC system. LC/MS combination is a method of choice for high throughput experiments and analysis of complicated biomedical samples.
Proteome analysis of spectra is made by Thermo Proteome Discovery software, that incorporates SEQUEST search engine.
Chromatography
ÄKTApurifier 100 core systems include Monitor UV-900 for UV-Vis detection of up to three wavelengths simultaneously. This allows visualization of more proteins when adequate detection at 280 nm cannot be achieved. Optional for ÄKTApurifier 100 core systems is Monitor pH/C-900, a combined monitor for accurate, online measurement of pH, conductivity, and temperature.
Gel electrophoresis
Our Core Facility has all necessary equipment for 2 dimensional gel electrophoresis. The detection of gels is performed by Bio-Rad ChemiDoc XRS System. We also have Agilent 2100 Bioanalyzer to run protein gel chips, and Agilent 2100 Bioanalyzer for FACS analysis on chips.
The Bio-Rad ChemiDoc XRS System is Bio-Rad’s most advanced imaging system. The system is able to image chemiluminescent, fluorescent and colorimetric samples and comes complete with a compact darkroom, transilluminator and supercooled high resolution 1.44 megapixel CCD camera. The entire system is controlled by the provided Quantity One 1-D analysis software with additional controls on the compact darkroom to control focus, aperture and zoom. Consequently, the ChemiDoc XRS system is extremely flexible and can be used for imaging a diverse range of applications such as Western blotting, DNA/RNA gel electrophoresis, 2-D gel electrophoresis, densitometry, etc. The system is able to detect HRP and AP chemiluminescence, ethidium bromide, SYBR® Green, green fluorescent protein, silver stain and Coomasie brilliant blue staining to name but a few that are frequently used with the above applications.
Bio-Rad ChemiDoc XRS System is equiped with 2D gel analysis software.
Sergei Nekhai, Ph.D.