Matrix-Assisted Laser Desorption Ionization MALDI

In another report, an open-access channel (i.e., no cover plate) has been used for CE separation before the MALDI-MS analysis [792]. 

One of the requirements in MALDI-MS analysis is the use of a liquid matrix. The electrowetting-on-dielectric (EWOD) method has been used to move and mix droplets containing proteins and peptides with the liquid matrix, all of which were situated at specific locations on an array of electrodes. With this method, insulin (1.75 pM), insulin chain B (2 pM), cytochrome c (1.85 pM), and myoglobin (1.45 pM) have been analyzed [518]. 

FIGURE 7.40 Penetration of a 10-pm-thick aluminum foil by a side-opened monolithic Si microneedle. Note that no damage can be observed on the needle [791]. Reprinted with permission from the Institute of Physics Publishing. 

FIGURE 7.41 Picture of the microfabricated fluidic device integrated with a standard MALDI-TOF sample plate. Because of the self-activating character of the microfluidic device, the system can be introduced into the MALDI ionization chamber without any wire or tube for the sample introduction and the flow control [820]. Reprinted with permission from the American Chemical Society. 

The research in the area of on-chip MS analysis is very active and the important differences are summarized in Table 7.1. 

Matrix-Assisted Laser Desorption Ionization (MALDI) 

Mele and Malpezzi [51] observed, that in the case of methylated CyDs abundant peaks are recorded in MALDI spectra corresponding to CyD adducts with DHB and metal cations (Na , K ). No such peaks are present for native CyDs. This observation indicates that the MALDI technique should be used with caution in the analysis of CyDs because of possible clustering of the matrix with the analyte. 

Interestingly, MALDI/TOF was also used for studying self-assembled mono- 

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A connnon feature of all mass spectrometers is the need to generate ions. Over the years a variety of ion sources have been developed. The physical chemistry and chemical physics communities have generally worked on gaseous and/or relatively volatile samples and thus have relied extensively on the two traditional ionization methods, electron ionization (El) and photoionization (PI). Other ionization sources, developed principally for analytical work, have recently started to be used in physical chemistry research. These include fast-atom bombardment (FAB), matrix-assisted laser desorption ionization (MALDI) and electrospray ionization (ES). 

Until about the 1990s, visible light played little intrinsic part in the development of mainstream mass spectrometry for analysis, but, more recently, lasers have become very important as ionization and ablation sources, particularly for polar organic substances (matrix-assisted laser desorption ionization, MALDI) and intractable solids (isotope analysis), respectively. 

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. 

Ionization can be improved in many cases by placing the sample in a matrix formed from sinapic acid, nicotinic acid, or other materials. This variant of laser desorption is known as matrix-assisted laser desorption ionization (MALDI). The vaporized acids transfer protons to sample molecules (M) to produce protonated ions [M + H]+. 

The ablated vapors constitute an aerosol that can be examined using a secondary ionization source. Thus, passing the aerosol into a plasma torch provides an excellent means of ionization, and by such methods isotope patterns or ratios are readily measurable from otherwise intractable materials such as bone or ceramics. If the sample examined is dissolved as a solid solution in a matrix, the rapid expansion of the matrix, often an organic acid, covolatilizes the entrained sample. Proton transfer from the matrix occurs to give protonated molecular ions of the sample. Normally thermally unstable, polar biomolecules such as proteins give good yields of protonated ions. This is the basis of matrix-assisted laser desorption ionization (MALDI). 

Laser-desorption mass spectrometry (LDMS) or matrix-assisted laser desorption ionization (MALDI) coupled to a time-of-flight analyzer produces protonated or deprotonated molecular ion clusters for peptides and proteins up to masses of several thousand. 

Matrix-assisted laser desorption/ionization (MALDI) is widely used for the detection of organic molecules. One of the limitations of the method is a strong matrix background in low-mass (up to 500-700 Da) range. In present work an alternative approach based on the application of rough matrix-less surfaces and known as surface-assisted laser desoi ption/ionization (SALDI), has been applied. 

Electron impact (El), or Efectrospray ionization (ESI), or Matrix-assisted laser desorption ionization (MALDI)... 

With the identities and amounts of amino acids known, the peptide is sequenced to find out in what order the amino acids are linked together. Much peptide sequencing is now done by mass spectrometry, using either electrospray ionization (ESI) or matrix-assisted laser desorption ionization (MALDI) linked to a time-of-flight (TOF) mass analyzer, as described in Section 12.4. Also in common use is a chemical method of peptide sequencing called the Edman degradation. 

Mass spectroscopy is a useful technique for the characterization of dendrimers because it can be used to determine relative molar mass. Also, from the fragmentation pattern, the details of the monomer assembly in the branches can be confirmed. A variety of mass spectroscopic techniques have been used for this, including electron impact, fast atom bombardment and matrix-assisted laser desorption ionization (MALDI) mass spectroscopy. 

Matrix-assisted laser desorption ionization (MALDI) is not yet a technique that has been used extensively for LC-MS applications. It is included here because it often provides analytical information complementary to that obtained from LC-MS with electrospray ionization, as illustrated later in Chapter 5. 

Experimentation showed that the protein was not glycosylated and that the sequence at the iV-amino acid terminus corresponded to that expected. The C-terminus sequence, however, did not correspond to that predicted and these data were interpreted in terms of the presence of a heterogeneous, truncated, protein. A study of the tryptic digest fragments from this protein with matrix-assisted laser desorption ionization (MALDI) with post-source decay enabled the authors to suggest the positions at which the parent protein had been truncated. 

The unseparated digest mixture was studied directly by mass spectrometry using matrix-assisted laser desorption ionization (MALDI) and this showed six of the polypeptides detected by LC-MS and three of the expected polypeptides that had not been detected by LC-MS. In contrast, MALDI did not show three polypeptides observed by LC-MS. 

Matrix-assisted laser desorption ionization (MALDI) A method used for the ionization of high-molecular-weight compounds. In this approach, the analyte is crystallized with a solid matrix and then bombarded with a laser of a frequency which is absorbed by the matrix material. 

Wigge, P. A., Jensen, O. N., Holmes, S., Soues, S., Mann, M., and Kilmartin, J. V. (1998). Analysis of the Saccharomyces spindle pole by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. J. Cell Biol. 141, 967-977. 

The focus of this chapter is the development of a technique often called wholecell matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) or whole-cell MALDI-TOF MS. Some groups prefer to use terms such as intact or unprocessed rather than whole, but the intended meaning is the same regardless of which word is used. As noted in the first chapter of this book, there are many different methods for the analysis of bacteria. However, for the analysis of intact or unprocessed bacteria, whole-cell MALDI-TOF MS is the most commonly used approach. This method is very rapid. MALDI-TOF MS analysis of whole cells takes only minutes because the samples can be analyzed directly after collection from a bacterial culture suspension. Direct MALDI MS analysis of fungi or viruses is similar in approach1,2 but is not covered in this chapter. MALDI-TOF MS of whole cells was developed with very rapid identification or differentiation of bacteria in mind. The name (whole cell) should not be taken to imply that the cells are literally intact or whole. Rather, it should be taken to mean that the cells that have not been treated or processed in any way specifically for the removal or isolation of any cellular components from any others. In whole-cell analysis the cells have been manipulated only as necessary to... 

The introduction and eventual commercialization of matrix-assisted laser desorption/ionization (MALDI) and electrospray (ESI) allowed biomarker status to be extended to proteins in 1996.15"17 With a few exceptions, ESI has been used in conjunction with extractions and high-pressure liquid chromatography (UPLC) interfaced with mass spectrometry. MALDI, on the other hand, has been widely adapted for rapid analysis of intact organisms, supported by bioinformatics.1819... 

It is therefore not surprising that the interest in PyMS as a typing tool diminished at the turn of the twenty-first century and hence why taxonomists have turned to MS-based methods that use soft ionization methods such as electrospray ionization (ESI-MS) and matrix-assisted laser desorption ionization (MALDI MS). These methods generate information-rich spectra of metabolites and proteins, and because the molecular ion is seen, the potential for biomarker discovery is being realized. The analyses of ESI-MS and MALDI-MS data will still need chemometric methods, and it is hoped that researchers in these areas can look back and learn from the many PyMS studies where machine learning was absolutely necessary to turn the complex pyrolysis MS data into knowledge of bacterial identities. 

The ability to resolve and characterize complicated protein mixtures by the combination of 2DLC and online mass spectrometry permits the combination of sample fractionation/simplification, top-down protein mass information, and bottom-up peptide level studies. In our lab, the simplified fractions generated by 2D(IEX-RP)LC are digested and analyzed using common peptide-level analysis approaches, including peptide mass fingerprinting (Henzel et al., 1993 Mann et al., 1993), matrix-assisted laser desorption/ionization (MALDI) QTOF MS/MS (Millea et al., 2006), and various capillary LC/MS/MS methodologies (e.g., Ducret et al., 1998). 

In a separate study, a protocol for Matrix-assisted laser desorption-ionization (MALDI) imaging mass spectrometry (IMS) has been proposed.18 This IMS technique provides a new approach to visualize spatial distribution of thousands of molecular species, including peptides, proteins, and their metabolites in two- or three-dimensional levels. This approach may also provide a straightforward method of determining the tissue distribution of multiple peptides or proteins in a quantitative manner.18 Chu et al.19 reported a nondestructive molecular extraction method to obtain proteins from a single FFPE or frozen tissue section, without destroying the tissue morphology, such... 

The growing interest for the identification and characterization of polar and large compounds caused the development and the introduction of new ionization techniques, such as electrospray ionization (ESI)[4], and matrix assisted laser desorption ionization (MALDI),[5] and their more recent improvements, thus establishing new MS based approaches for studying large molecules, polymers and biopolymers, such as proteins, carbohydrates, nucleic acids. 

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