MS Bioworks differential analysis services (MSB-09, 10, 11, 12) are used to provide a quantitative analysis of two or more samples. This service can be used with or without fractionation. The extent of fractionation determines the number of proteins identified in a sample and the coverage obtained per protein. For the line items described here protein level fractionation is performed using SDS-PAGE. Alternative fractionation strategies such as reverse phase, Hydrophilic Interaction Liquid Chromatography (HILIC), anion exchange and cation exchange and removal of abundant proteins are available upon request. Label free (spectral counting and dMS) and Labeling strategies (SILAC and ITRAQ) are available. SILAC and ITRAQ approaches enable global proteome quantitation and are ideal for studying the impact of genetic or chemical modifications to a system.
The approaches described here are compatible with multiple sample types, for example biofluids, tissue, cell lines and spent media. MS Bioworks recommends submitting 20µg protein per sample. The table below represents a typical number of protein identifications from a cell lysate with different degrees of fractionation.
|Number of fractions||0||10||20||40|
|No. of Proteins (≥2 peptides/protein)||790||2521||4137||5093|
In the example presented below MCF10A cells were treated with the AKT kinase inhibitor A-443654 in a SILAC experiment. A global proteomic analysis was performed at the four hour time point. The levels of cathepsin B, a cysteine protease known to be involved in cancer progression and invasion, decreased fivefold as a result of drug treatment.
A total of 5093 proteins were identified in the MCF10A experiment. The data have provided a novel insight into the mechanism of A-443654.
MSB-09,10,11,12 can be ordered directly from our online ordering system, here.