Abstract
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 with 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 R~f~ 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. Published in 2008 by John Wiley & Sons, Ltd.