HPTLC/DESI-MS Imaging of Tryptic Protein Digests Separated in Two Dimensions
- ORNL
- Merck Research Laboratories
Desorption electrospray ionization mass spectrometry (DESI-MS) was demonstrated as a method to detect and identify peptides from two-dimensional separations of cytochrome c and myoglobin tryptic digests on ProteoChrom HPTLC Cellulose sheets. Data-dependent tandem mass spectra were acquired during lane scans across the TLC plates. Peptides and the corresponding proteins were identified using a protein database search software. Two-dimensional distributions of identified peptides were mapped for each separated protein digest. Sequence coverages for cytochrome c and myoglobin were 81% and 74%, respectively. These compared well to those determined using the more standard HPLC/ESI-MS/MS approach (89% and 84%, respectively). Preliminary results show that use of more sensitive instrumentation has the potential for improved detection of peptides with low Rf values and improvement in sequence coverage. However, less multiple charging and more sodiation were seen in HPTLC/DESI-MS spectra relative to HPLC/ESI-MS spectra, which can affect peptide identification by MS/MS. Methods to increase multiple charging and reduce the extent of sodiation are currently under investigation.
- Research Organization:
- Oak Ridge National Laboratory (ORNL)
- Sponsoring Organization:
- ORNL work for others; SC USDOE - Office of Science (SC)
- DOE Contract Number:
- AC05-00OR22725
- OSTI ID:
- 961557
- Journal Information:
- Journal of Mass Spectrometry, Journal Name: Journal of Mass Spectrometry Journal Issue: 12 Vol. 43; ISSN 1076-5174
- Country of Publication:
- United States
- Language:
- English
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