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Intact Protein MS Protocols
General considerations
  • Protein/peptide samples should be fully solubilized. Centrifuge and/or filter if necessary
  • Protein samples should be 10 ┬ÁM or less than 0.1 mg/mL
  • Protein samples should be free of detergents (no SDS, NP-40 or Tween-20 etc.)
  • Use an appropriate low salt buffer (avoid PBS or other non-volatile salts if possible; limit salts to <10 mM total)
  • No strong acids/bases: if possible avoid TFA (but no more than 0.1 %), use formic acid instead
  • No DMSO
Intact Protein LC-MS
  • For large intact proteins (>30 kDa), select a size exclusion column (SEC) method
  • For smaller proteins (MW 5-20 kDa), select a C4 column method
  • For peptides (MW <4 kDa), select a C18 method
  • A gradient of 0.1 % formic acid:acetonitrile is a good initial solvent system to try
In-gel Digest