ABSTRACT

MALDI (matrix-assisted laser desorption/ionization) IMS (imaging mass spectrometry) can provide the spatial distribution, relative abundance, and molecular identity of thousands of endogenous analytes of biological species (lipids, metabolites, peptides, proteins, and drugs) directly from tissue sections.(1-3) Mass spectra can be acquired at multiple x-y coordinates across a thin tissue section, and the intensities of mass-to-charge (m/z) values are used to reconstruct two-dimensional (2D) ion density maps. IMS is currently used in clinical and medical research where it has been successfully applied to the molecular analysis of diseases such as cancer and to aid in the assessment of clinical diagnosis and prognosis.(4-7) This technology has also been utilized to study normal biological processes to understand underlying molecular mechanisms. For instance, proteomic and lipidomic molecular maps have been generated to study early developmental stages of mouse embryo implantation.(8,9) Additionally, IMS has been used to determine the spatial distribution of exogenous pharmaceuticals and metabolites in animal organs and whole rat tissue sections.(10-12)

13.1 Introduction ................................................................................................269 13.2 The Tissue Imaging Experiment ................................................................ 271

13.2.1 Sample Preparation ...................................................................... 271 13.2.2 Mass Spectrometric Analysis ...................................................... 272

13.3 Applications of Tissue Imaging ................................................................. 274 13.4 Applications of Tissue Imaging by Ion Mobility-Mass Spectrometry ...... 277 13.5 Pharmaceutical Applications ......................................................................280 13.6 Conclusions and Perspectives ..................................................................... 282 References .............................................................................................................. 282

In the past decade, advances in MS instrumentation have allowed more effective analysis of tissue sections by MALDI IMS. In particular, the addition of a rapid postionization gas-phase ion mobility separation allows an ion fractionation process to be integrated into the imaging experiment. MALDI imaging ion mobility MS separates analytes first by collision cross section (CCS) in the mobility cell and second by m/z in the MS analyzer.(13-17) The ion separation step before MS analysis provides a rapid (μs-ms) separation of complex samples without additional sample preparation or a significant increase in analysis time. For the imaging experiment in particular, the additional ion fractionation step provides the capability of resolving two isobaric analytes by structure alone, producing independent images.(18) Ion mobility can separate different classes of biomolecules based on their intrinsic gas-phase packing efficiencies along CCS versus m/z trend-lines.(19) The point at which a given analyte falls along these trend-lines provides qualitative information of the molecular species without the need for MS/MS analysis (Figure 13.1). As visualized in a 2D conformation plot in Figure 13.1a, lipids have larger CCS (Å2) values than peptides of the same m/z. A benefit of the ion mobility separation for the analysis of complex samples, such as tissue sections, is illustrated in Figure 13.1b where peptides are separated and exported apart from the more abundant endogenous phospholipids. Ion mobility separation additionally enhances the IMS experiment by separating analytes of interest from endogenous chemical noise and provides the capability of selectively analyzing or imaging one class of biomolecules in the presence of another. Ion mobility can also be used to target peptides containing post-translational modifications that deviate from the peptide trend-line and simultaneously fragment all species after ion mobility separation, which is especially applicable to high-throughput pharmaceutical imaging.(18,20-22)

FIGURE 13.1 MALDI ion mobility MS analysis of a grade II human astrocytoma tissue section. (a) The 2D conformation plot with predicted phospholipid and peptide trend-lines are indicated by dashed lines. Ion mobility MS signal intensity is indicated by false coloring (scale displayed). (b) Signals from the phospholipid and peptide trend-lines were exported separately and plotted as m/z to intensity.