Matrix-assisted laser desorption ionization imaging mass spectrometry: in situ molecular mapping

Abstract

Matrix-assisted laser desorption ionization imaging mass spectrometry (IMS) is a relatively new imaging modality that allows mapping of a wide range of biomolecules within a thin tissue section. The technology uses a laser beam to directly desorb and ionize molecules from discrete locations on the tissue that are subsequently recorded in a mass spectrometer. IMS is distinguished by the ability to directly measure molecules in situ ranging from small metabolites to proteins, reporting hundreds to thousands of expression patterns from a single imaging experiment. This article reviews recent advances in IMS technology, applications, and experimental strategies that allow it to significantly aid in the discovery and understanding of molecular processes in biological and clinical samples.

Figure 1

Overview of an IMS experiment illustrated on a section of adult mouse heart. (A) A fresh frozen section is mounted on a conductive target surface with a serial section used for hematoxylin and eosin (H&E) stain. (B) The matrix is applied by sublimation, and the section is irradiated by the laser in an array pattern. (C) A mass spectrum is generated for each x,y coordinate. (D) Selected ions are then mapped across the tissue to create images. Images may be combined to form a multiplex ion image. Abbreviations: SM, sphingomyelin; PC, glycerophosphocholine. (E) Images may be combined to form a multiplex ion image.

Figure 2

Experimental workflow that should be considered when designing an IMS experiment.

Figure 3

SRM imaging of sinalbin on N. caerulescens on new to older leaflets. The matrix 9-aminoacridine (15 mg/mL in a 9:1 methanol/water mixture) was manually sprayed onto the plant leaf and imaged by MALDI LTQ XL operating in MS/MS mode at a spatial resolution of 100 μm. The image is from the m/z 424 → m/z 259 + 275 transition. Sinalbin was detected with a higher concentration in newer leaflets, in agreement with the known protective role of sinalbin in new leaflets. Contributed by M. Reyzer at Vanderbilt University (Nashville, TN) and D. H. McNear at the University of Kentucky (Lexington, KY).

Figure 4

Combined ion image from four lipids from an imaging experiment on a mouse embryo at embryonic day 13.5. The tissue section was washed with 50 mM ammonium formate, followed by sublimation with DHB. IMS data were collected at a 20 μm spatial resolution in negative ion mode. Lipids are assigned by mass using lipidmaps.com.

Figure 5

Peptide imaging illustrated on a portion of a tissue microarray (TMA). The TMA is treated with trypsin, after being mounted on a MALDI target. A pathologist marks an optical image of a serial section stained with H&E as cancer (outlined in red) or normal (outlined in black). This image is then overlaid on the optical image of the TMA prepared for IMS. The TMA is spotted with trypsin, and then matrix and is analyzed by IMS. Marked regions are then compared to formulate molecular signatures that differentiate tumor and normal tissue. A classification model is produced from these signatures for the analysis of additional samples. Here, regions of images that classified as normal are colored yellow, whereas the images that classified as cancer are colored red (image courtesy of E. Seeley and T. Morgan of Vanderbilt University).

Figure 6

Protein images from adult rat kidney tissue. Sinapinic acid was applied by sublimation followed by rehydration, and the tissue was imaged at a spatial resolution of 100 μm. Images are shown at 100% of that peak’s intensity normalized to total ion current. Image contributed by J. Yang of Vanderbilt University.