Parallel post-source decay for increasing protein identification confidence levels from 2-D gels.
Peptide mass fingerprinting (PMF) has over the years become one of the most commonly used tools for high-throughput analysis and identification of proteins. This method is applicable when relatively simple samples have to be analysed and it is commonly used for analysing proteins previously separated by 2-DE. The most common type of instrument used for this approach is the MALDI-TOF that has proved to be particularly suitable for the PMF analysis because of its characteristics of speed, robustness, sensitivity and automation. We have used a MALDI-TOF equipped with a novel parallel PSD capability (MALDI micro MX), to perform the analysis of two sets of different biological samples isolated by 2-DE. By using a method that integrates the data obtained by PMF analysis with the PSD data obtained in the same experiment, we show that the new multiplexed PSD solution increases the protein identification rate compared to the normal PMF approach. We also investigated the use of a charge-directed fragmentation modification reagent to improve the identification rate and confidence levels.
Wahlander
Asa
A
Department of Protein Technology, Lund University, Lund, Sweden.
Arrigoni
Giorgio
G
Snel
Marten
M
Hellman
Ulf
U
James
Peter
P
D000818:Animals; D002460:Cell Line; D030562:Databases, Protein; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D006801:Humans; D007668:Kidney / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008239:Lysine / Q000737:chemistry; D051379:Mice; D010449:Peptide Mapping / Q000295:instrumentation* / Q000379:methods*; D010455:Peptides / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry; D040901:Proteomics / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000295:instrumentation* / Q000379:methods*
D010455:Peptides; D011506:Proteins; D008239:Lysine