MALDI-MS Imaging

MALDI-MS, particularly with an atmospheric pressure MALDl source, is a powerful method that allows the analysis and detection of a wide variety of biomolecules including lipids directly from tissue sections. The laser serves two purposes (1) to desorb analytes from the surface material and (2) induce analyte ionization for MS analysis. The role of the matrix is to absorb the majority of the laser energy, leading to explosive desorption of the matrix crystals, together with incorporation of analyte into the gas phase without degradation of the analyte. Addition of matrix may also aid the ionization of analyte molecules in the gas phase due to the presence of adduct ions from the added modifier(s). 

During a MALDI-IMS experiment, the laser is fired across the surface of the matrix covered tissue section, which allows desorption and ionization of lipids. 

Nowadays, pulsed, frequency tripled Nd YAG UV lasers (355 nm) are usually employed for MALDI experiments with a repetition rate of 1000 Hz in commercial instruments for sufficient data acquisition. In MALDI-IMS, the resolving power for application strongly depends on the sample preparation step (e.g., matrix crystal size), stepper motor accuracy, and laser spot sizes. To achieve MALDI-IMS to a practical resolution, the laser spot size of 20 pm is usually used. Therefore, the time needed to obtain images from a sample depends on the number of analyzed spots, the repetition rate of the laser (Hz), and the data collecting and processing speed of computers. For example, imaging a whole-body mouse or rat section with current commercially available MALDI mass spectrometers equipped with lasers operating at 1 kHz would take 2-4 h. 

From IMS of lipids, alterations in lipid distribution due to changes of physiological or pathological conditions could be determined. For example, age-dependent changes in the distribution and amount of PC species in rat brain have been previously evaluated [57] and changes in localization of lipids after traumatic brain injury in rat brain have been determined [72]. A study by MALDI-IMS dynamically demonstrated the changes in skeletal muscle lipid composition induced by contraction [40]. The investigators found the reduction of DAG and TAG, as well as the accumulation of PC species in the contracted muscles. 

Studies with IMS of lipids can address many specific scientific questions related to the distributions of particular types of lipids. For instance, determination of the distribution of GPL speeies containing polyunsaturated fatty acids in a particular type of cells in a mouse brain section [57] of gangliosides in different regions of the brain [73] of ST in different layers of rat hippocampus [74] and of PC and GalCer species in rat brain sections [75] has all been well addressed. 

M S imaging is perceived as a rather new - but very promising - approach, not only for the basic sciences but also for clinical diagnosis [156]. Although other MS imaging methods based on desorption electrospray-ionization (DESI) MS [157] or secondaryion mas s spectrometry (SIMS)are currently available (for a timely review, see Ref. [158]), attention here will be focused exclusively on MALDI-MS imaging. The same technique is also discussed comprehensively in Chapter 4 of this book. 

In addition to brain (which is often investigated for the reasons indicated above), the lipid compositions of many different tissues have already been successfully investigated using MALDI-TOF-MS imaging [162], and the compositional data 

Unfortunately, the achievable resolution is normally not sufficient to obtain images with cellular resolutions [163]. Thus, a combination between MALDI and SIMS imaging represents a very promising approach since, whilst SIMS is capable of providing images with a subcellular resolution, MALDI is unequivocally the method of choice for detecting lipids with relatively high masses, such as cardioli-pins [164]. 

A comprehensive review of MS imaging, and not only of the related lipid aspects, is available in Chapter 4. 

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Professor Caprioli of Vanderbilt University (US), Professor Heeren s group (Europe), and our group (Japan) independently developed the MALDI-MS imaging and attained the expected resolution.1,2... 

Figure 12.5. (Top) MALDI-MS images of clozapine in rat brain slice as a function of laser fluence (bottom) relative MALDI responses of clozapine and norclozapine in rat brain slice as a function of laser fluence.

Figure 12.8. MALDI-MS images of letters (a) S and (b) P containing norclozapine and clozapine in rat brain sections, respectively. MALDI-MS/MS spectra of (c) clozapine and (d) norclozapine in rat brain sections (Hsieh et al., 2006a).

In addition, we also envisage MALDI-MS imaging becoming more mainstream in the area of carbohydrate research, offering the added dimension of spatial information on top of the specificity of the structural information that MS delivers. 

Beyond peptides and proteins, MALDI MS imaging of tissue section for the detection of low-molecular-weight compounds can also be achieved. Of particular interest is the posttreatment location of pharmaceutical compounds in targeted tissues or organs. Further, in parallel to location, the effects of a drug on the local proteome can be observed as a function of dose or time. Variations in the proteome are indicative of drug efficacy.116... 

Monroe E, Annangudi S, Hatcher N, Gutstein H, Rubakhin S, Sweedler J (2008) SIMS and MALDI MS imaging of the spinal cord. Proteomics 8 3746-3754. doi 10.1002/pmic.200800127... 

Goodwin R, Dungworth J, Cobb S, Pitt A (2008) Time-dependent evolution of tissue markers by MALDI-MS imaging. Proteomics 8 3801-3808. doi 10. 1002/pmic.2008002001... 

Jurchen J, Rubakhin S, Sweedler J (2005) MALDI-MS imaging of features smaller than the size of the laser beam. J Am Soc Mass Spectrom 6 1654-1659. doi 10.1016/j. jasms.2005.06.006... 

The acquisition of the mass spectra for peptide and protein imaging is done in our lab on commercial MALDI TOF instruments (Voyager sSTR, 4700 Proteomics Analyzer Applied Biosystems, Framingham, MA) equipped with a Nd YAG laser. The acquisition of the data is controlled by an in-house designed software (MALDI MS Imaging Tool, MMSIT). The area to be scanned is specified as a pattern of points, equidistant from each other. The distance is chosen referring to the diameter of the... 

FIGURE 11.2 General procedure for MALDI-MS imaging analysis and conventional quantitative HPLC/MS analysis. 

FIGURE 11.7 (a) MALDI-MS images of clozapine in rat brain slice as a function of laser... 

During the past several years, significant progress was made for the MALDI-MS imaging technique. It has been proven to be a valuable tool for studying drug distribution in animal tissues. MALDI-IMS is a simple, quick, and molecularly specific analytical platform that can distinguish small molecules from their metabolites on... 

Ohkawa, T. and Bunch, J., MALDI MS imaging to reveal distribution of benzodiazepine drug and metabolite molecules in rat brain, Proceedings of the 55th ASMS Conference, Indianapolis, IN, 2007. 

Whole-body MALDI MS imaging of proteins has also been performed and an example is presented in Fig. 1.3. Images from an embryonic day 14.5 mouse are shown one stained and optically imaged and the other imaged by MALDI MS. In the latter, each of the five colors shown represents a different protein (unpublished data courtesy Peggi Angel, Vanderbilt University). 

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