Delayed extraction time-of-flight MALDI mass spectrometry of proteins above 25,000 Da.
Matrix-assisted laser desorption/ionization (MALDI) with delayed extraction (DE) has been optimized for mass analysis of high-mass proteins and glycoproteins with masses above 25,000 Da. Under optimized experimental conditions, i.e. using a weak extraction field strength and a long delay time, a steep drop in mass resolution above 30,000 Da is no longer observed and an improvement of more than a factor of 10 is obtained compared with the non-DE case, at least up to 66 kDa in a 1.2 m time-of-flight mass analyzer. On this level of resolution the factors limiting further improvements become apparent, i.e. adduct ion formation between matrix and analyte, but also cationization and further non-matrix-related adducts, as well as prompt fragmentation. Moreover, heterogeneity of the sample is often the reason for the detection of broad signals for larger proteins. Within these limitations, DHBs (a 9:1 mixture of 2,5-dihydroxybenzoic acid and 2-hydroxy-5-methoxybenzoic acid) gave by far the best results.
Bahr
U
U
Division of Instrumental Analytical Chemistry, J. W. Goethe University of Frankfurt, Germany.
Stahl-Zeng
J
J
Gleitsmann
E
E
Karas
M
M
D006023:Glycoproteins / Q000032:analysis; D008970:Molecular Weight; D011506:Proteins / Q000032:analysis / Q000302:isolation & purification*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D006023:Glycoproteins; D011506:Proteins