Suport

All users should use these sample preparation guidelines in order to improve the quality of the results. If you have any questions, do not hesitate to contact the Mass Spectrometry Center

 

Mass Spectrometry of small molecules

- Mass spectrometry is a powerful analytical tool.  However, impurities and contaminants are especially important as signal intensity and spectra quality depends mainly on the quality of samples. Thus, proper sample preparation is one of the keys to maintain a successful mass spectrometric assay.
- It is important that you can estimate the amount of sample submitted for MS analysis. From a practical point of view, for ESI experiments, we prefer that you submit ~1 mg. However this is not required.  
- Check the solubility the compound you are submitting for MS analysis.  Solvents commonly used in Mass Spectrometry are dichloromethane, methanol, acetonitrile and water. If a preferred solvent is not mentioned, Methanol will be used as solvent for MS analysis.
- ESI and MALDI tolerate a limited concentration of salts. The presence of salts or surfactants can suppress the signal for other components. If your sample contains salts, contact the Mass Spectrometry Center. Samples with phosphate salts can not be analyzed be ESI-MS.

 

Proteomic services

-Our proteomics service includes in-gel and/or in-solution proteolytic digestion, MALDI-TOF (MS and MS/MS) for the identification of proteins based on database searching. Under request our services may include peptide separation by liquid chromatography before MS analysis.
- By routine for in-gel proteins identification, samples submitted as gel slices are de-stained using appropriate protocols and  proteins are  digested  with trypsin overnight (other enzymes can be used, please contact the Mass Spectrometry Center). Under request, reduction and alkylation can also be used for in–gel digestion.
- Data obtained is searched against predicted fragments from the trypsin digestion of proteins contained in a database (Sprot, NCBI, etc.) using Mascot.
- We encourage our clients to use Coomassie or colloidal blue stain. Other stains are possible but please contact first the Mass Spectrometry Center for advice.
- Please note that extreme caution has to be used to avoid contamination with keratin, especially for low level protein sample.
- To avoid contamination with keratin wear powder free latex gloves all the time when processing the samples. Loose hair should be secured back. All trays and equipment must be rinsed with water before used.

Samples can be submitted:

In-gel: an excised bend from a gel. The band should be cut using extreme caution to avoid keratin contamination. A band that can be visualized by Coomassie stain, usually contains sufficient protein for identification.

In solution: samples can be submitted in solution (10-20 l). Avoid any detergents and unnecessary sample buffers.

Pre-digested samples: a pre-digested sample can be submitted for analysis by using the protocols used in the Mass Spectrometry Center.

 

-Sample submission procedure-

Gel samples submission:

  1. Minimize keratin contamination.

Remember to work in a clean air environment, clean the bench; always use powder free latex gloves, brand new pipette tips, razor blades and tubes

  1. Gel bands should be ~1 x 5mm; gel spots should be ~2mm diameter.
  2. Place in 1.7mL eppendorf tube covering gel pieces with 0.01% acetic acid.
  3. Specify source organism and fill our request service form.

Protein solution sample submission:

  1. Buffer composition.

The solution background should be water or < 50mM volatile salt (e.g. TEAB) and when possible avoid detergents, surfactants and non-volatile salts.

  1. Ship proteins refrigerated or lyophilized.
  2. Specify source organism and fill our request service form.