Proteomics Core FacilityProtocols

Gel staining prior to protein identification

We accept samples for protein identification from coomassie and silverstained gels. Coomassie staining is preferred, so only use silverstaining if the amount of protein is insufficient for visualization by coomassie.

Coomassie staining

Essentially all coomassie stains are compatible with mass spectrometry, including GelCode Blue Stain (Pierce), Bio-Safe Coomassie stain (Bio-Rad) and the Colloidal Blue Staining kit from Invitrogen.
A protocol for colloidal coomassie staining can be downloaded here.

Silver staining

Only some silver staining protocols are compatible with mass spectrometry. Do NOT use commercial kits, we only accept samples that have been stained using this protocol.

Cutting of gel bands

If you have gels from which you like individual bands to be identified by MS, you have two options. First, you can send (or bring) the gel to the facility, and we cut out the band(s) that you indicate.

Alternatively, you can cut out gel bands yourself. In this second scenario, please take precautions to prevent contamination (keratins, etc).

Please use the following guidelines.  

Mass determination of intact proteins

The molecular mass of proteins is best determined under denaturing conditions. Please observe the following guidelines for sample preparation:

• Provide at least 10 ul of a protein at a concentration of >1 mg/ml
• Avoid the use of detergents

Only in very specific cases we can analyse proteins under non-denaturing (or native) conditions.

• Provide at least 100 ul of a protein at a concentration of >2 mg/ml
• The protein must be in a volatile buffer, preferably 25 mM ammoniumacetate at any pH below 7. Plain 25 mM ammoniumacetate pH = 6.7, for pH adjustment use acetic acid.
• Avoid the use of detergents