Endogenous peptides from biophysical and biochemical fractionation of serum analyzed by matrix-assisted laser desorption/ionization and electrospray ionization hybrid quadrupole time-of-flight.
Blood peptides can be concentrated, extracted, and analyzed with strong signal-to-noise ratios by precipitation in organic solvents followed by extraction in water. Matrix-assisted laser desorption/ionization (MALDI) and electrospray ionization (ESI) hybrid quadrupole time-of-flight (Qq-TOF) were used to analyze the precipitated and extracted endogenous peptides from fetal calf serum. C18 solid-phase extraction with or without prior precipitation in ammonium sulfate, size exclusion chromatography, dealbuminization, dye affinity chromatography, ultrafiltration, and differential precipitation in organic solvents were compared. Hundreds of different ions could be observed by MALDI in the various fractions. It appeared that some peptides were freely dissolved and that not all peptides in blood were obliged to remain bound to albumin or other high-molecular-mass proteins. Mass spectra with high signal-to-noise ratios were obtained from polypeptides precipitated with organic solvents followed by extraction of the peptides from the pellet with water. The peptides extracted from organic precipitates were analyzed by nano liquid chromatography (LC)-ESI-Qq-TOF. In addition to many commonly abundant serum proteins, apparent low-abundance peptides associated with cancer biology from proteins such as insulin-like growth factor II, thymosin beta4 and beta9, plasminogen, coagulation factors, and extracellular matrix protein 1 were observed.
Tucholska
Monika
M
Department of Chemistry and Biology, Faculty of Engineering and Applied Science, Ryerson University, Toronto, Ont., Canada M5B 2K3.
Scozzaro
Salvatore
S
Williams
Declan
D
Ackloo
Suzanne
S
Lock
Chris
C
Siu
K W Michael
KW
Evans
Kenneth R
KR
Marshall
John G
JG
D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000737:chemistry; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods; D008969:Molecular Sequence Data; D010455:Peptides / Q000097:blood* / Q000302:isolation & purification; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D001798:Blood Proteins; D010455:Peptides