UCHSC Proteomics and Mass Spectrometry Core Facility

Preparing Samples for MALDI

Best results are obtained from clean samples (buffers, salts, detergents.. interfere with MALDI)

Contaminating species are poorly tolerated in MALDI and will require removal if small, uniform matrix/analyte crystals do not form.
Ziptip clean up works well for most protein (C4) and peptide (C18) applications.
If you have a sample that needs to be reconstituted use ultrapure water, acetonitrile is acceptable to use if needed for solubility.
We will add trifluoroacetic acid (TFA) to acidify the sample.

Preparing Samples for LC-ESI

Best results are obtained from clean samples (buffers, salts, detergents.. interfere with ESI)

Upfront reverse phase liquid chromatography allows for analysis of peptide and protein samples with salts (~< 100 mM) but in most cases not detergents, or polymers.
If you have a sample that needs to be reconstituted use ultrapure water, do not use organic solvents.
We will add formic acid (FA) to acidify the sample. TFA will reduce the sensitivity of positive ion mode by approximately 10 fold.

Definitions (with Respect to Mass Spectrometry)

Resolution = mass/full width at half height

Parts Per Million (ppm)- Refers to mass accuracy = Dmass (theoretical to measured) x 10^6/mass (measured)

example: measured mass: 999.99 Da theoretical mass: 1000.00 Da error: 10 ppm

Mass Spectrometry Terms Project

Posttranslational Modifications

Posttranslational modifications (PTM) can often be identified by MS (although it is rarely a trivial task). More starting material is required compared to identification experiments. There is no guarantee that a modified peptide will ionize readily, in fact many modifications decrease the ionization efficiency of a peptide. For a review see Larsen et. al. Biotechniques 40:790-798, 2006.

Biotechniques 40:790-798, 2006