Peptides & Proteins

Concentration & Volume

  • For routine intact mass analysis, the minimum amount of sample depends on the MW of the peptide or protein. Good results have been obtained with 25 pmol at 5 kD, 100 pmol at 20 kD, 200 pmol at 40 kD, and 500 pmol at 60 kD.
  • Sample concentration should be such that the appropriate amount of protein is contained in 20-25 ul.

Salts & Buffers

  • Concentrations of non-volatile components (e.g. NaCl, Na2HPO4 etc.) should be kept below 10mM, if not eliminated completely.
  • Concentrations of Glycerol, DMSO or DMF should be kept under 1%.
  • Detergents like Tween, Triton etc. should be avoided, since they give strong signals in MS.
  • Volatile buffers such as ammonium acetate and formate are tolerable below 20 mM.
  • In case of ESI measurements, samples should not contain TFA (e.g. from a reverse-phase HPLC run); therefore, TFA has to be removed by lyophilization (don´t use Speedvac).
  • The salt and buffer limits become increasingly stringent as the MW of the protein increases. To maximize the likelihood of a successful analysis, proteins greater than 20 kD should ideally be submitted in deionized water only.