Gel Electrophoresis

The cell or tissue proteome is a highly complex mixture of tens or even hundreds of thousands of proteins. Direct analysis of such a complex mixture to identify changes in protein expression associated with the onset of disease first requires some form of separation of the individual components. The best established and most exquisite form of protein separation is 2DE whereby proteins are separated from each other based on their charge and size. Unlike traditional techniques which use a single wide pH range for the first dimension, Proteome Sciences has developed a platform based on the use of multiple narrow pH ranges to increase the number and quality of protein spots visible on the final 2DE gel. With this and other proprietary technology, the focus has been to increase protein detection to fentomolar levels (10^-15) and beyond.

Whilst the serum proteome is less complex than that of cells, its analysis is complicated by the presence of a small number of highly abundant proteins. These mask the less abundant proteins when using 2DE and so they must be removed prior to analysis.

Proteome Sciences uses commercially available depletion products and proprietary methods for depletion in serum and other sample types and applications.

In addition to improvements in the performance of its 2DE system, Proteome Sciences uses software packages that link the unique sample identification number to all stages of the 2DE proteomics analysis. The software contains a number of analytical modules which can process gel images from diseased and control samples and identify the protein spots which are different between them. This can be further fine-tuned by the scientist to produce a map of regulated proteins. The software then interacts with automated gel-cutters which excise the regulated protein spots from the 2D gel and process them for mass spectrometry. The software collects data from the mass spectrometry analysis for each regulated spot and links this to the protein identification results that are obtained through data mining both public and proprietary databases.

Proteome Sciences’ laboratories are equipped to perform high throughput proteomics with the capacity to run over 300 2DE gel protein maps per week. The automated gel-cutters can each process 1,150 spots per day, allowing over 11,000 differentially regulated proteins to be processed by mass spectrometry per week. The excised spots are analysed using a combination of LC-MS, LC-MS/MS on MALDI-TOF and ESI-TOF/TOF mass spectrometers. For protein identification Proteome Sciences has access to all the public protein sequence databases.

HIGH OUTPUT AND HIGH THROUGHPUT CAPABILITY