Protein separation

State-of-the-art equipment for the quest for the protein of interest

ÄKTA purifier 10

The ÄKTA purifier 10 is a versatile liquid chromatography device, suitable for different kinds of protein and peptide purification, including size-exclusion chromatography, affinity chromatography, ion exchange chromatography or reversed phase chromatography – depending on the desired end-result. Monitoring is possible by UV detection (280nm or 254 nm) and conductivity measurements. The included fraction collector enables the user to automatically collect all required fractions of variable volumes.

Gel-based proteomics

Our gel-based proteomics platform is built up with BioRad-equipment and accessories.

Criterion Vertical Electrophoresis Cell

For 1D SDS-PAGE, e.g. precast TGX gels (Tris-Hcl based) are run in a Criterion Vertical Electrophoresis Cell. This electrophoretic device is intended for running gels of midi-format of different buffer systems. The used polyacrylamide gels vary from 12+2-wells (loading volume max. 45 ul), to 18 (30 ul) and 26 (15 ul) wells. A standard gel run with protein samples dissolved in classic Laemmli buffer loaded on a TGX gel takes about 1 hour (200 V).

Protean IEF Cell

For 2D protein separation, an extra isoelectric focusing step is performed, prior to the SDS-PAGE. ReadyStrip IPG strips are run in a Protean IEF Cell for protein separation based on their isoelectric point. Different pH gradients for these strips are possible, depending on the requested application.

Trans-blot Cell

When gels are further used for Western blotting, the protein transfer is done in a Trans-Blot Cell, in which up to 6 midi-gels can be run simultaneously. Different buffers are possible for this tank blotting system; in general, transfer is done with CAPS buffer.

Protein identification and separation

From broad screening to detailed (quantitative) information

In 2014, ProGenTomics made the strategic choice of pursuing the innovative "Data-independent Acquisition" (DIA) methodologies. Two LC-MS platforms were specifically developed for doing exactly that at the time: The TripleTOF 5600 (Sciex) and the Synapt G2Si (Waters) QTOF instruments. Have a closer look at what these Mass Spectrometers can do:

TripleTOF 5600 (Sciex)

The Triple TOF 5600 is coupled to a microflow LC system (Eksigent LC425). While still not common ground, microflow LC has gained significant sensitivity over the past years and can now provide unprecedented stability and thus quantitative accuracy at the cost of only 2x loss in sensitivity. This is of particular importance when running SWATH runs. Herein, a library of peptide identifications is first generated on all the samples in the experiment using conventional DDA acquisition. Next, these same samples are acquired by sequential fragmentation of small m/z windows to generate quantitative information on the transitions found in the library. It is, in essence, a post-acquisition MRM-like data-extraction approach providing the user with a permanent record of everything. This implies that future hypotheses can be pursued in a targeted analysis long after the samples were acquired using SWATH.

No need to say that the intrinsic sensitivity and speed of this instrument equally allow for very high quality DDA runs as well. For specific purposes, e.g. iTRAQ labelling might be required and this would be the preferred platform to do such analysis.

SynaptG2Si (Waters)

The Synapt G2Si is a very versatile Q-TOF device comprising Ion Mobility separation (IMS) and Electron Capture Dissociation (ETD) capabilities. It allows for a plethora of different acquisition strategies, the most prominent of which is HDMSE or ion mobility-assisted data-independent acquisition. Herein, both identification and quantitation is done on DIA data. In ProGenTomics, this instrument is coupled to a Nano Acquity 2D LC (RPRP) system providing very high sensitivity: whole proteome analyses by HDMSE are done on 100ng of protein extract per sample.

About ProGenTomics

ProGenTomics, located in Gent (Belgium), offers a wide range of proteomics and genomics services and support to the academic community and its external customers. We aim to tailor our services to the needs of our collaborators and customers, while giving great support.  

Contact us

ProGenTomics, proteomics & genomics services

  Ghent University, Faculty of Pharmaceutical Sciences

  Ottergemsesteenweg 460, 9000 Gent, Belgium

 

  +32 9 264 83 56