The goal is to determine which proteins are present in your sample. The sample can be a cellular extract, a gel spot or band, a purified protein, etc.
We typically digest the sample with trypsin after reduction of disulfide bonds and alkylation of free thiol groups. This can be performed in solution or directly in gel.
The resulting peptides are analyzed either by LC-MS/MS (most of the samples) or by MALDI-TOF/TOF (mostly 2D gel spots, or high-throughput identification of pure proteins). Once data recorded, they are submitted to database search using Mascot search engine in order to identify proteins. Amino acid sequences from the organism of origin have therefore to be available in public databases like SwissProt, NCBI, etc. or should be provided to be inserted in an in-house confidential database.
What is delivered?
A list of proteins identified with relevant statistical values. For these proteins, you will know which peptides were identified.
Report and results will be sent by email.
What is needed for these analyses?
- For in solution digestion, at least 10 µg of proteins are required. Please note that if the buffer is not compatible with the workflow, an additional purification step will be performed
- For in gel digestion, the bands/spots must be visible by Coomassie blue staining to be identified.
- For 1D-PAGE: please provide us the entire gel. The picking will be performed manually at the facility. Beyond 10 bands to pick, a fee will be charged per band to excise.
- For 2D-PAGE/DIGE: gel spots must be 1.5 mm in diameter and be transferred in an Eppendorf twin.tec PCR plate 96, semi skirted (order no: 0030 128.575) and wells A2, A4, A6, A8, A10 and A12 have to be empty on each 96 wells plate.
What is the time to get results?
Results are generally available in 10 working days