Applications and performance of a MALDI-ToF mass spectrometer with quadratic field reflectron technology.
A new matrix-assisted laser desorption/ionization time of flight mass spectrometer (MALDI-ToF MS), developed specifically for the identification and characterization of proteins and peptides in proteomic investigations, is described. The mass spectrometer which can be integrated with the 2-D gel electrophoresis workflow is a bench-top instrument, enabling rapid, reliable and unattended protein identification in low-, as well as high-throughput proteomics applications. To obtain precise information on peptide sequences, the instrument utilizes a timed ion gate and a unique quadratic field reflectron (Z2 technology), allowing single-run, post-source decay (PSD) of selected peptides. In this study, the performance of the instrument in reflectron, PSD and linear mode, respectively, was investigated. The results showed that the limit of detection for a single peptide in reflectron mode was 125 amol with a signal to noise ratio exceeding 20. Average mass resolution for peptides larger than 2000 u was around 13,000 full width, half maximum (FWHM). The limit for protein identification during peptide mass fingerprinting (PMF) was 500 amol with a sequence coverage of 18%. Mass error during PMF analysis was less than 15 ppm for 17 out of 25 (68%) identified peptides. In PSD mode, a complete series of y-ions of a CAF-derivatized peptide could be obtained from 3.75 fmol of material. The average mass error of PSD-generated fragments was less than 0.14 u. Finally, in linear mode, intact proteins with molecular masses greater than 300,000 u were detected with mass errors below 0.2%.
Flensburg
John
J
Amersham Biosciences AB, Björkgatan 30, SE-751 84 Uppsala, Sweden. john.flensburg@amersham.com
Haid
Daniel
D
Blomberg
Johan
J
Bielawski
Jacek
J
Ivansson
Daniel
D
D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D010455:Peptides; D011506:Proteins