Sample submissions for Electrospray (ESI) Or Matrix-Assisted Laser Desorption Ionization (MALDI) MS Analysis are normally required as purified materials without any salt. Proteins are dialyzed or digested in either ammonium bicarbonate solution (<50 mM) or 10mM Tris-HCl buffer. Samples containing SDS detergent or glycerol are not suitable for MS analysis. If high concentration salts are not avoidable, we recommend the use of on-line LC-ESI MS or off-line LC–MALDI MS analysis.
The Coomassie blue R250 or colloidal Coomassie blue (i.e. Pierre Gel Code®) are the preferred protein stains for identification of 1D or 2D PAGE resolved bands. In-gel digestion of silver-stained or SYPRO Ruby stained proteins often yield low sequence coverage even if the same amount of protein was applied. If you are unable to obtain Coomassie Blue stainable amounts of protein you must use a mass spectrometry compatible silver staining protocol (see silver staining protocol below).
Keratins are the common contaminates from in-gel digests, which cause a big problem in the mass spectrometric analysis of low abundance proteins, especially for silver-stained gels. We strongly suggest that gels should be treated very careful to avoid such impurities from human skin, hairs, and trypsin.