Sigma Xi Postdoctoal Poster Presentation Abstract
Presenter name: Matthew J. Vergne
Mentor name: Michael J. Welch
Room and Building: Room A153, Building 227
Mail Stop: 8392
Sigma Xi member: no
Mass Spectrometry Studies on the Reproducibility of Protein Digestion
Matthew J. Vergne, David M. Bunk, and Michael J. Welch
Analytical Chemistry Division, Chemical Science and Technology Laboratory
National Institute of Standards and Technology, Gaithersburg, MD 20899-8392
Mass spectrometry quantification of individual proteins from biological samples has the potential to be a more selective and accurate method than immunoassays routinely performed in clinical chemistry. However, mass spectrometric quantification of intact proteins has proven to be difficult due to structural heterogeneity of macromolecular protein. A different approach to quantification is to determine the amounts of protein fragments (peptides) produced from trypsin digests with the assumption that the molar amount of each studied peptide is equal the molar amount of the studied protein. To determine the feasibility of peptide quantification, the reproducibility and completeness of protein digests was tested.
Bovine serum albumin (BSA) was chosen as a model protein. Three BSA in-solution trypsin digests were prepared on three different days for a total of nine digests. Each sample was digested over a 48-hour time period. The digest solutions were analyzed using capillary LC/MS. The peak area ratios of peptide pairs with similar elution times in each sample were calculated. The coefficients of variation (CV) of the area ratios between samples and between days were evaluated along with the precision of the method itself. The feasibility of quantification of peptides using isotopically labeled internal standards of peptides will be evaluated.