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This service is a quantitative and qualitative analysis of two or more complex samples such as cell lysates, tissue homogenates or biofluids. The results from this experiment provide a catalogue of the proteins present in all samples and a statistical analysis reflecting the changes in the protein levels observed across the samples, along with associated pathway information.
Samples may be fractionated prior to analysis; the extent of fractionation performed dictates the number of proteins observed. The greater the number of fractions, the greater the number of proteins identified and sequence coverage of those proteins.
A 10 fraction analysis is suitable for the analysis of low complexity samples e.g. HPLC fractions, semi-purified extracts. A 20 fraction analysis is for medium complexity samples e.g. subcellular fractions, pooled HPLC fractions. A 40 fraction analysis is suitable for complex samples e.g. whole cell lysates, whole tissue extracts.
Samples or fractions are digested using trypsin and the peptide mixture is analyzed by LC/MS/MS. The peptides are fragmented in the mass spectrometer to yield diagnostic patterns that can be matched to protein sequence databases via computer algorithms.
A minimum of two samples must be submitted for differential analysis.
1. Sample types: Lysates, cells, tissues, biofluids, FFPE protein extracts, SILAC
2. Relative quantitation of protein abundance can be carried out without any biological or chemical labeling of proteins or peptides (label-free quantitation) or by labeling proteins metabolically with amino acids incorporating stable isotopes (e.g. SILAC) or by labeling proteolytic peptides using chemical tags that incorporate stable isotopes (e.g. ICAT, iTRAQ). We use the Scaffold® software for label-free quantitation and the Mascot® or ProteinPilot® software for quantitation of samples labeled with isotopic or isobaric tags.
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