MALDI matrix application

Another factor that determines spatial resolution for MALDI—MSI is the size of the matrix crystals formed during the matrix deposition process. The size of the sample matrix co-crystals grown is strongly dependent on the sample matrix solution composition and the rate at which the crystals are grown (Cohen and Chait, 1996). For the majority of MALDI—MSI experiments, the spot size of the laser is such that multiple crystals are sampled in each laser shot, thus the spatial resolution is limited by the laser spot size and not the crystals formed. However, it is still important to avoid nonuniformities in the matrix layer (crystals), which can cause ionization yields to vary across the sample and hinder the interpretation of spatial information. Some approaches for MALDI matrix application, such as inkjet printing (Nakanishi et ah, 2005 Baluya et ah, 2007), can produce a uniform coating of small crystals. Different approaches for the application of MALDI matrix will be discussed further in Section 14.4.4.2. 

Tissues were incubated with RNase, then rinsed 10 min with successive SSC solutions and twice 0.5xSSC solutions at 55°C for 30 min. After rinsing slices with O.lxSSC for 5 min at room temperature, one bath of ultrapure water was carried out to remove the excess of polymers. Tissues were kept drying at room temperature before MALDI matrix application. 

Typical for arthropod (such as insects, spiders, crustaceans) neurosecretory tissue is that the secretory part of the glands is the outside surface see Fig. 25.1b 1), which, therefore, is readily accessible for MALDI matrix application. Furthermore, the tiny size of the tissue allows for whole mount tissue analysis (i.e., without the need of a cryostat) and for full gland analysis within a relatively short time frame. An additional advantage is that live... 

MALDI Matrix Application Dried Droplet Versus Spray... 

The second method of sample preparation for IMS is a matrix-coating method for MALDI imaging. In this chapter, we review the choices of matrix compound and solvent composition appropriate for IMS of tissue sections. Three kinds of matrix-application methods and examples of their use are illustrated. 

Also for MALDI, there is a special case worth mentioning. Surface-enhanced laser desorption/ionization (SELDI) is a technique that utilizes special sample plates [196, 197]. These have different modified surfaces, for example, hydrophobic, anionic, or antibody treated. Which type of surface to select depends on the application. After application of analyte the surface is washed according to a protocol leaving only the desired components on the target. Finally, a MALDI matrix is applied before analysis in the spectrometer. See Chapter 12 for an application example of SELDI. 

Although CHCA works well with most drug molecules, there is no universal MALDI matrix. However, as with the internal standard, it is not practical to optimize matrix selection for each compound in broad drug discovery applications. 

The synthesis of phosphopeptides is typically confirmed mass spectrometri-cally using either a MALDI (matrix-assisted laser desorption/ionizafion) or an ESI (electrospray ionization) source, and the peptide purity is determined by reversed-phase chromatography coupled to an UV detector (Figs. 1 and 2B). Whenever possible, phosphopeptide analyses should be complimented by mass spectra recorded in negative ion mode. For most biochemical applications it is necessary to purify the peptides by HPLC techniques (Fig. 2A). 

FIGURE 11.1 (See color insert following page 210.) General overview of MALDI-IMS. (a) Fresh section cut from sample tissue, (b) Mounted section after matrix application using a robotic picoliter volume spotter, (c) Partial series of mass spectra collected along one row of coordinates U-axis). (d) Three-dimensional volumetric plot of complete dataset with selected m/z slices or ion images. Principal axes are x, y, and m/z. Color of each voxel is determined by ion intensity. (From Cornett, D. et al., Nat. Methods, 4, 828, 2007. With permission.)... 

Apart from automating the matrix application process, it is critical to evaluate the resulting matrix crystals and coating for analyte extraction, localization, and effect on tissue architecture. Several reports have shown that even for standard analytes mixed with matrices, there is an uneven distribution of the analyte within the MALDI crystals and that sample preparation influences the resulting distribution [18-20], Thus, other active areas of research are focused on optimizing matrix application and sample preparation protocols. 

During the last decade, processing of polymers has become an important field of applied and fundamental research [48]. One of the most important fields is laser ablation involving various techniques and applications. Laser ablation is used as an analytical tool for MALDI (matrix-assisted laser de-sorption/ionization) [28, 29] and LIBS (laser-induced breakdown spectroscopy) [49] or as a preparative tool for PLD (pulsed laser deposition) of inorganic materials [37] and of synthetic polymer films [50, 51]. Another application is surface modification of polymers [52] if low fluences are applied, the polymer surface can be either chemically modified to improve adhesion... 

Methods relying on MALDI require application of an extraction solvent to the layer to move sample to the layer surface followed by co-crystallization with a MALDI matrix material. TLC-MALDI direct coupling methodologies use one of three methods of layer... 

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