Small & Macromolecule Protocols | Mass Spectrometry | SIU

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Mass Spectrometry Facility

Small & Macromolecule Protocols

Sample Preparation for MS (Must Read!)

Below you will find a number of guidelines that should be followed when preparing a sample for mass spectrometric analysis.

  • All samples should be submitted in polypropylene microcentrifuge tubes/vials (for peptides or proteins samples) or deactivated glass vials with Teflon-lined screw caps (for small molecule samples). Use a small volume insert if a limited amount of sample is available.
  • All sample tubes must be labeled with a sample ID and be accompanied by a sample submission form.

  • For MALDI analysis, please submit at least 1mg of purified solid sample or 50uL of (at least) 10pmol/mL sample solution prepared in compatible solvent(s) or a volatile buffer.

  • For electrospray analysis, please submit 50uL of (at least) 10pmol/mL sample solution prepared in 50% acetonitrile.

  • Solvents that are compatible with MALDI include acetonitrile, methanol, isopropanol, ethanol and water.

  • A volatile buffer such as ammonium acetate should be used whenever pH control is necessary. The maximum concentration that can be tolerated is 50 mM.

  • Allowable detergents include n-octyl-glucoside, n-dodecyl-glucoside, octanoyl-N-methylglucamide and decanoyl-n-methylglucamide

  • Avoid using:

    • Tris, phosphates, citrates, and HEPES

    • Urea and guanidinium salts

    • TFA (acts as strong ion-pairing agent in electrospray)

    • Detergents such as SDS, CHAPS, PEG, Tween, and Triton

    • DMSO, acetates and  glycerol

    The presence of any of the above chemicals can completely suppress ionization of the sample making the difference between a successful or failed analysis.

Protein Staining (Compatible with MS)

Coomassie Staining Silver Staining

In-Solution Tryptic Digestion 

In-Gel Tryptic Digestion

Coomassie Stained (1D) Coomassie Stained (2D)

Silver Stained (1D)Silver Stained (2D)

Reverse Phase C18  Cleanup and Concentration for Peptides