Matrix-coated tissue section

Place the ITO slide with the matrix-coated tissue section into the MALDI adapter plate. 

Imaging involves collecting MS data in a regular pattern or array across a matrix-coated tissue section (Fig. IB). This approach typically provides a higher resolution molecular picture of the tissue from data collected from hundreds or thousands of pixels across the tissue surface. An image displaying the distribution... 

Most recently, Caprioli s group183 have reported a novel method of dry coating tissue sections with MALDI matrix, thus minimizing the problem of the matrix solvent mobilizing the surface analytes. The dry coating procedure proved to be simple and rapid and yielded high-quality images of phospholipids. 

Application of a matrix to tissue sections is a critical step in MALDI imaging. As mentioned above, the target analytes should not change their in vivo position and should not spread or smear during use of the MALDI matrix. The following procedures have been developed to achieve a uniform coating of a matrix ... 

Figure 8.2 Design of protein-embedding barcode is depicted in (a) five thin layers of matrix (the thicker lines) coated with variable concentration of tested protein (thinner lines located above the matrix), (b) A FFPE tissue section of bladder cancer IHC-stained by monoclonal antibody to E-cadherine showing variable intensity of positive staining results which is compared with a protein-embedding bar code as designed in this chapter. Using computer-assisted image analysis with a special software, an automatic quantitative measurement of protein is performed. See color insert.

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. 

One can adjust the thickness of the matrix coat covering a tissue section by varying the number of cycles of coating and drying. 

Han and Schey [71] used a special matrix coating method in processing the bovine lenses sections (30 10 pm) for MS imaging. The tissue sections were first sprayed with an acetonitrile-water (50 50, vol/vol) solution resulting in a tightly bound section. After drying, the tissue sections were coated with a thin layer matrix of S A at 15 mg/mL in ethanol-water (50 50, vol/vol). After it was dried, the tissue sections were finally sprayed with several cycles of SA matrix solution at 15 mg/mL in ethanol-water-formic acid (44 44 12, vol/vol/vol). 

For a few tissue samples, the matrix deposition can be performed manually. A matrix solution is sprayed onto the tissue section with a hand-held thin layer chromatography (TLC) sprayer or an artist airbrush. The reproducibility of manual matrix deposition is an issue. When the manual sprayer is used, the MALDI target plate with the tissue section is held vertically about 15-25 cm from the sprayer nozzle. It is recommended to spray multiple coats of matrix across the tissue section and each coating cycle consists of passing the sprayer two to hve times across the tissue section and allowing the tissue to dry for about 1-5 min. This process is usually repeated between 10 and 20 cycles. 

A TLC reagent sprayer/or a glass spray nebulizer was used for the matrix coating onto tissue section by several researchers [36, 44, 76, 122, 133], Puolitaival et al. [134] reported a solvent free matrix Dry-Coating method. The matrix (DHB) was ground for 15 min using a mortar and pestle into crystals between 1 and 40 pm in... 

Astemizole [152] examined the spatial distribution of astemizole and its metabolites in rat brain slices with and without perfusion with saline solution. The Sprague-Dawley rats were treated orally with the drug at 100 mg/kg in 0.4 % methylcellu-lose. Matrix solution (DHB, 10 ml) coated by 15-20 coats over the entire surface of tissue sections by a glass reagent sprayer. MALDI-MS/MS images showed the distribution of astemizole and its metabolite (M-14) in rat brain slice. Astemizole appeared to be the major drug-related component in rat brain (Fig. 2). 

Since its inception about 15 year ago, MALDI-IMS has been developed into a powerful and versatile tool for biomedical research. It allows for the investigation of the spatial distribution of molecules at complex surfaces. The combination of molecular speciation with local analysis makes a chemical microscope that can be used for the direct biomolecular characterization of histological tissue section surface. However, successful detection of the analytes of interest at the desired spatial resolution requires careful attention to several steps in the IMS protocol matrix selection, matrix coating, data acquisition, and data processing. MALDI-IMS is increasingly playing an important role in the drug discovery and development and disease treatment. 

In addition to frozen tissue sections, tissue blotting and laser-capture microdisection methodologies can also be used. In the former method, the proteins are transferred by blotting freshly dissected tissues onto an active Cig-coated surface. Another surface of choice for blotting is a carbon-filled polyethylene membrane. In the laser-capture microdisection method, a specific population of cells from a stained tissue is transferred onto an ethylene-vinyl acetate transfer film. A matrix is applied to the isolated cells through a narrow capillary. 

FIGURE 3.2 Schematic representations of MALDI-IMS procedures. Usually, the tissue section mounted on an indium thin oxide (ITO)-coated glass shde is covered with a specific MALDI matrix. Next, the ITO slide is inserted into a mass spectrometer. The MALDI laser scans through a set of preselected locations on the tissue (10-200 pm scan pitch) and the mass spectrometer records the spatial distribution of molecular species. Suitable image processing software can be used to import data from the mass spectrometer to allow visualization and comparison with the histological image of the sample. 

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