MALDI profiling

The seeds peptide MALDI profile proved to be suitable in characterizing grape varieties. This result was confirmed excluding, or by evaluating as being of minor influence, the other factors that might contribute to the protein profile the harvest year, zone of production and vineyard treatments. To achieve it, seeds of Raboso Piave grape samples collected from different vineyards and harvested in different years were studied. The 3D plot of Fig. 10.3 shows that the profile is maintained in particular the species at m/z 6113, which is characteristic of this variety, is detected in the spectra of all samples. 

Figure 10.3. Peptide MALDI profiles of Raboso Piave seed extracts of grapes harvested from two different vineyards in two different years (Pesavento et al., 2008).

Figure 15.7. Experimental protocol for MALDI profiling and imaging of mammalian tissue samples. (From ref. 19.)...

Lasers can be used in either pulsed or continuous mode to desorb material from a sample, which can then be examined as such or mixed or dissolved in a matrix. The desorbed (ablated) material contains few or sometimes even no ions, and a second ionization step is frequently needed to improve the yield of ions. The most common methods of providing the second ionization use MALDI to give protonated molecular ions or a plasma torch to give atomic ions for isotope ratio measurement. By adjusting the laser s focus and power, laser desorption can be used for either depth or surface profiling. 

Some solid materials are very intractable to analysis by standard methods and cannot be easily vaporized or dissolved in common solvents. Glass, bone, dried paint, and archaeological samples are common examples. These materials would now be examined by laser ablation, a technique that produces an aerosol of particulate matter. The laser can be used in its defocused mode for surface profiling or in its focused mode for depth profiling. Interestingly, lasers can be used to vaporize even thermally labile materials through use of the matrix-assisted laser desorption ionization (MALDI) method variant. 

MALDI-ToF is a technique that allows the molecular weights of proteins and peptides to be determined. It is less susceptible to suppression effects than electrospray ionization and thus is able to be used for the direct analysis of mixtures. In the case of a crude tryptic digest, MALDI-ToF will provide a molecular weight profile of the polypeptides present without the analysis time being extended by the need to use some form of chromatographic separation. 

Figure 5.19 MALDI-ToF mass spectrum, providing a molecular-weight profile of the tryptic peptides derived from spot 22 (see Figure 5.18) of the silver-stained two-dimensional gel of the proteins extracted from the yeast S. cerevisiae. From Poutanen, M., Salusjarvi, L., Ruohonen, L., Penttila, M. and KaUddnen, N., Rapid Commun. Mass Spectrom., 15, 1685-1692, copyright 2001. John Wiley Sons Limited. Reproduced with permission.

Advances in size-exclusion chromatography, coupled with refractive index, absorption, viscosity, and lightscattering detectors, and MALDI-ToFMS, have made it possible to accurately determine molecular weight distribution (oligomer profiling), even at the relatively low values of polymeric additives (up to about 5000 Da). Advances in column design, e.g. high-resolution PS/DVB columns (> 105 plates m-1) mean that SEC can provide a valuable alternative to conventional HPLC techniques for the separation of small molecules. 

C. Tempst, P Serum peptide profiling by magnetic particle assisted automated sample processing and MALDI-TOF mass spectrometry. Anal. Chem. 2004, 76, 1560-1570. 

Holland et al. proposed a simpler and faster approach using the protein profile obtained directly from whole cells rather than from cellular extracts or affinity beads.17 This was the first report of the whole-cell MALDI-TOF MS... 

An interesting variation on the whole-cell MALDI approach was recently reported in a study aimed more at FTMS than TOF MS, but the results are nevertheless interesting and important to users of both methods for analysis of bacteria 40. Wilkins s group showed both MALDI-TOF and MALDI-FTMS spectra of whole bacteria grown on isotopic media depleted in C13 and N14. Because most bacterial identification protocols involve a culture step prior to analysis, it is possible to manipulate the sample based on control of the growth media. For mass spectral analysis manipulation of the isotope profile... 

Approximately 90% to 95% of whole-cell MALDI MS profiles are representative of the ribosomal proteins abundant in rapid growth whole cells. Although the identification of proteins, from whole bacterial cells, by MALDI-TOF MS analysis is ambiguous, at best, due to the low-mass accuracy and resolving power, several researchers realized that many of the observed... 

The results for bacterial whole-cell analysis described here establish the utility of MALDI-FTMS for mass spectral analysis of whole-cell bacteria and (potentially) more complex single-celled organisms. The use of MALDI-measured accurate mass values combined with mass defect plots is rapid, accurate, and simpler in sample preparation then conventional liquid chromatographic methods for bacterial lipid analysis. Intact cell MALDI-FTMS bacterial lipid characterization complements the use of proteomics profiling by mass spectrometry because it relies on accurate mass measurements of chemical species that are not subject to posttranslational modification or proteolytic degradation. 

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