Ionization MALDI

The hybrid can be used with El, Cl, FI, FD, LSIMS, APCI, ES, and MALDI ionization/inlet systems. The nature of the hybrid leads to high sensitivity in both MS and MS/MS modes, and there is rapid switching between the two. The combination is particularly useful for biochemical and environmental analyses because of its high sensitivity and the ease of obtaining MS/MS structural information from very small amounts of material. The structural information can be controlled by operating the gas cell at high or low collision energies. 

MALDI ionization represents an ideal method for directly sampling the most abundant and basic proteins directly from whole cells. Typically these are ribosomal proteins. TOF mass spectrometry can be easily coupled to MALDI... 

Selection of an appropriate internal standard can also assist in correcting for ion suppression issues caused by matrix components. However, if the ion suppression is too severe, then inevitably sensitivity will suffer. Ion suppression must be limited to a degree sufficient to avoid sensitivity problems. To reduce ion suppression, a sample cleanup method is necessary. Moreover, proper co-crystallization is directly related to sample composition and therefore, sample cleanup is necessary for successful MALDI ionization. 

Knochenmuss, R., and Zenobi, R. (2003). MALDI ionization The role of in-plume processes. Chem. Rev. 103 441-452. 

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. 

MALDI differs from ESI in two major aspects (a) the MALDI ionization process is not continuous, but is a pulsed event, and (b) while the analytes for ESI are dissolved in the appropriate solvent and introduced into the ionization source, the MALDI analytes are dissolved and mixed with a solution of the appropriate matrix and then are cocrystallized with the matrix. Sensitivity for both techniques enables analyses at the low femtomole range (and even below), although most analyses are conducted at the low picomole range because of the greater convenience of sample handling at this level. Mass measurement accuracy, as indicated elsewhere in this discussion, depends on the choice of analyzer. These techniques are complementary and neither has exhibited dominance in the analysis of biological molecules. 

Preferably, electrospray ionization (ESI) is used in combination with quadrupole mass filters [16,17], whereas MALDI is commonly used in combination with the time-of-flight (TOF) analyzer [18], The relatively simple construction of these two types of analyzers and the resulting price advantage has led to their replacing the traditional magnetic sector instruments as the workhorses of mass spectrometric analysis. A more recent development is the ion-cyclotron-resonance (ICR) analyzer [19] which can be used for both ES-and MALDI-ionization. 

The ES/MALDI-FT-ICR mass spectrometer of the Institute of Organic Chemistry at the University of Tubingen is from Bruker Daltonik GmbH, Bremen. The system is evacuated by efficient turbo molecular pumps which allows HPLC and GC coupling over long time periods. ES, NanoSpray and MALDI ionization sources allow individual adaptation to particular problems. Samples from combinatorial chemistry can be routinely analyzed with the 60°-ESI from Analytica of Branford Inc. (Branford, MA) via LC/ES-FT-ICR-MS as well as small amounts of valuable biological samples with nanoESI. 

Figure 16.6 A simplified schematic of a time of flight spectrometer and the principle of the ion reflector (reflectron). (1) sample and sample holder (2) MALDI ionization device by pulsed laser bombardment (3 and (3 ) ions are formed between a repeUer plate and an extraction grid (PD 5000V) then accelerated by an other grid (4) control grid (5) microchannel collector plate (6) signal output. Below, a reflectron, which is essentially an electrostatic mirror that is used to time-focus ions of the same mass but which have initially different energies. The widths of the peaks are of the order of 10 and the resolution ranges between 15 to 20 000.

MALDI ionization mostly takes place in vacuum in spite of the recent development of atmospheric pressure ionization sources. Therefore, coupling a microfluidic system to MALDI-MS implies working under vacuum on the chip or performing the MS analysis off-line, i.e. introducing the microchip in the MALDI-MS once the fluidic operations are finished. Consequently, the first miniaturized developments for MALDI-MS analysis only concerned the... 

In this book we endeavor to cover most of the topics highlighted in the introduction, and to present (i) technological developments achieved for the coupling of microfluidic systems to MS, both for the ESI and MALDI ionization techniques, (ii) relevant applications of microfluidics-to-MS couplings and (iii) research aiming at miniaturizing the mass spectrometer itself. We hope thereby to give a complete and comprehensive view to the reader of the miniaturization and mass spectrometry field with this collection of chapters. 

A schematic of a MALDI-TOF-MS instrument is depicted in Figure 11.2b. Samples, consisting of a few microlitres of analyte solution (with or without matrix), are deposited on a MALDI target (Figure 11.2a). After the solvent has evaporated the sample plate, carrying the solidified samples, is introduced into the MALDI ionization chamber via load-lock. The ionization process takes place in a high-vacuum chamber to which the plate is introduced via a prechamber kept at a pressure lower than atmospheric. Analyte ions are then accelerated as they are formed and pumped into the TOF analyzer, where they are separated based on their mass-to-charge ratio. 

The book covers several subtopics of miniaturization and mass spectrometry. It combines (i) technological developments in the quest for miniaturization of sample preparation and how to connect micrometer-sized devices to a mass spectrometer, (ii) various illustrations of fields that benefit from such a hyphenated technique and (iii) technological developments for the miniaturization of the mass spectrometer. Additionally, the book is not restricted to one ionization technique as is often the case for many reviews, but it reports on efforts for both the ESI and MALDI ionization techniques. After an introduction to miniaturization and mass spectrometry, the book is divided in three sections that respectively concern (i) ESI-MS applications, (ii) MALDI-MS applications and... 

A major breakthrough in the analysis of proteins and peptides came about with the development of sensitive methods based on the use of mass spectrometry. The importance of these developments was recognised in 2002 with award of the Nobel Prize in Chemistry to John Fenn (electrospray (ESI) ionization) and Koichi Tanaka (matrix-assisted laser desorption/ionization (MALDI) ionization) pioneers in the development of methods of ionisation that made protein and peptide MS a practicable procedure. These developments have led to MS being the method of choice for protein identification and characterisation of protein and peptides (Fig. 

The data shown in Figure 9.5 are from a mixture of sphingomyelin (ML), dyn-orphin (MP), and chlorisondamine (Chi). The primary feature of this 2D spectrum is the presence of two families of ions. Because this spectrum was obtained with a MALDI ionization source, all the ions are singly charged. So, unlike the ion families discussed for Figure 9.3, these trend lines do not arise from multiple charges but from structure similarities among compound classes. 

Ions can be formed within the vacuum chamber of the mass spectrometer or outside the instrument at atmospheric pressure. Examples of in vacuo ionization are electron ionization (El), chemical ionization (Cl), and matrix-assisted laser desorption/ion-ization (MALDI). Ionization techniques carried out outside the vacuum system are collectively termed atmospheric pressure ionization (API). The most important API methods are electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI). Among other API techniques are atmospheric pressure photo-ionization... 

How Analyte Ions Are Formed Several diverse views have been expressed on this topic [33-38]. No single model provides a complete picture of MALDI ionization. The widely accepted current view is that the analyte ionization is a two-step process a primary ionization event, followed by in-plume secondary ion-molecule reactions [34-36]. In the first step, reactive matrix ionic species are generated. The analyte ions are produced in the expanding gas plume (an area just above the matrix surface) via extensive secondary ion-molecule charge-transfer reactions between the primary matrix ions and neutral analyte molecules ... 

J. J. Corr, et al., Design considerations for high-speed quantitative mass spectrometry with MALDI ionization, J. Am. Soc. Mass Spectrom. 17, 1129-1141 (2006). 

MALDI ionization is mainly performed in the vacuum region of the mass spectrometer however, it can also be performed at atmospheric pressure (Moyer and Cotter, 2002 Schneider et al., 2005). One of the major advantages of MALDI is that in can be adapted very easily, using the atmospheric pressure interface on virtually any type of mass spectrometer. 

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