Figure 1
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Scientists often select the newest generation of innovative matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF)
and MALDI-TOF-TOF mass spectrometers to take advantage of their extraordinary research capabilities. For example, an important
MALDI-TOF and MALDI-TOF-TOF application is high-success protein identification from 2D gels. Here, MALDI-TOF-TOF is the foremost
component of new proteomics analysis systems for peptide mass fingerprinting, peptide fragment analysis, and posttranslational
modifications (PTM) analysis. The newest generation of ultrahigh-performance TOF-TOF mass spectrometers enables high-throughput
protein identification by MALDI-TOF peptide mass fingerprinting, immediately followed by more detailed, high-caliber protein
characterization using MALDI-TOF-TOF tandem mass spectrometry (MS-MS) on the same prepared sample. Comprehensive MS-MS information
is delivered to TOF-TOF users from rather minute sample amounts within just a few seconds. In addition, new T3-sequencing
capabilities allow top-down sequence analysis on even many intact proteins (1).
The newest generation of top TOF-TOF instrumentation provides scientists with two MS-MS methods for de novo sequencing. For
highest specificity, laser-induced decomposition with post-fragmentation acceleration TOF-TOF MS is the method of choice,
providing clear spectra with readily available amino acid and sequence-specific ions such as i-, a-, b- and y-ions. For in-depth analysis of selected peptides, high-energy collision-induced dissociation (CID) is very valuable as well.
Regarding protein identification and quantitation by LC-MALDI, new advanced approaches provide unique, intelligent LC workflow
for LC–MALDI experiments. With new isotope coded protein label (ICPL, Toplab GmbH, Martinsried, Germany) triplex technology
now available, three independent entire proteomes can be quantitated even simultaneously. Low-abundance proteins are identified
and can be characterized in detail using MALDI-TOF-TOF, in which identifying and characterizing low-abundance proteins are
considered the "holy grail" of proteomics.
Figure 2
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Sophisticated workflows further enhance results through a synergistic data-dependent combination with LC–electrospray ionization
(ESI)-MS-MS data. LC–ESI-MALDI-MS-MS wizard-driven workflows utilize complementary information from the two ionization techniques
in a single eclectic experiment, providing much more useful in-depth knowledge and a new level of insight. Quantitative expression
proteomics experiments with various isotope-labeling techniques including ICPL, isotope coded affinity tag (ICAT, University
of Washington, Seattle, Washington), and isotope tagging for relative and absolute protein quantitation (iTRAQ, Applera Corp.,
Norwalk, Connecticut) are now being realized as well. Flexible quantitation software is becoming future proofed, allowing
scientists to incorporate new or unique isotope labels.
Meanwhile, many major developments have been taking place concerning advanced biomarker panel discovery, validation, and identification.
Here again, MALDI-TOF-TOF plays a commanding role, providing seamless integration into clinical proteomics solutions for high-resolution
peptide and protein putative biomarker panel discovery, identification, and validation. Generally, such solutions are approved
for research only, and are not yet in the clinic — the next big step.
