A recent innovation in mass spectrometry is the ability to record mass spectra on ordinary samples, in their native environment, without sample preparation or preseparation by creating ions outside the instrument. In desorption electrospray ionization (DESI), the principal method described here, electrically charged droplets are directed at the ambient object of interest; they release ions from the surface, which are then vacuumed through the air into a conventional mass spectrometer. Extremely rapid analysis is coupled with high sensitivity and high chemical specificity. These characteristics are advantageously applied to high-throughput metabolomics, explosives detection, natural products discovery, and biological tissue imaging, among other applications. Future possible uses of DESI for in vivo clinical analysis and its adaptation to portable mass spectrometers are described.

https://science.sciencemag.org/content/sci/311/5767/1566.full.pdf

Ambient Mass Spectrometry

1and has been initially targeted for the analysis of peptides and proteins present on or near the surface of tissue sections 2 . Imaging MS brings a new tool to bear on the problem of unraveling and understanding the molecular complexities of cells. It joins techniques such as immunochemistry and fluorescence microscopy for the study of the spatial arrangement of molecules within biological tissues. Many previous experiments using MS to image samples have focused on the measurement of the distribution of elements and small molecules in biological specimens, including tissue slices and individual cells 3‐5 . An extensive review on imaging by MS can be found in the article by Pacholski and Winograd 6

Imaging mass spectrometry: a new technology for the analysis of protein expression in mammalian tissues.

Imaging mass spectrometry combines the chemical specificity and parallel detection of mass spectrometry with microscopic imaging capabilities. The ability to simultaneously obtain images from all analytes detected, from atomic to macromolecular ions, allows the analyst to probe the chemical organization of a sample and to correlate this with physical features. The sensitivity of the ionization step, sample preparation, the spatial resolution, and the speed of the technique are all important parameters that affect the type of information obtained. Recently, significant progress has been made in each of these steps for both secondary ion mass spectrometry (SIMS) and matrix-assisted laser desorption/ionization (MALDI) imaging of biological samples. Examples demonstrating localization of proteins in tumors, a reduction of lamellar phospholipids in the region binding two single celled organisms, and sub-cellular distributions of several biomolecules have all contributed to an increasing upsurge in interest in imaging mass spectrometry. Here we review many of the instrumental developments and methodological approaches responsible for this increased interest, compare and contrast the information provided by SIMS and MALDI imaging, and discuss future possibilities.

Imaging mass spectrometry

Analytical characteristics of DESI are summarized. Examples of applications to small and large molecules, to in situ analysis, and to high-throughput analyses are presented. Evidence is provided for both a heterogeneous charge-transfer mechanism and a droplet pick-up mechanism of ionization. The speed, lack of the need for sample preparation, selectivity, and sensitivity of DESI are all demonstrated and discussed. Instrumentation is also discussed. Forensic applications as well as emerging areas of application including tissue imaging are given emphasis.

https://www.onlinelibrary.wiley.com/doi/pdf/10.1002/jms.922

Ambient mass spectrometry using desorption electrospray ionization (DESI): instrumentation, mechanisms and applications in forensics, chemistry, and biology

Matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry (IMS) is emerging as a powerful tool for investigating the distribution of molecules within biological systems through the direct analysis of thin tissue sections. Unique among imaging methods, MALDI-IMS can determine the distribution of hundreds of unknown compounds in a single measurement. We discuss the current state of the art of MALDI-IMS along with some recent applications and technological developments that illustrate not only its current capabilities but also the future potential of the technique to provide a better understanding of the underlying molecular mechanisms of biological processes.

/pdf/maldi-imaging-mass-spectrometry-molecular-snapshots-of-54ybmt728y.pdf

MALDI imaging mass spectrometry: molecular snapshots of biochemical systems.

Matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) has been used to generate ion images of samples in one or more mass-to-charge (m/z) values, providing the capability of mapping specific molecules to two-dimensional coordinates of the original sample. The high sensitivity of the technique (low-femtomole to attomole levels for proteins and peptides) allows the study of organized biochemical processes occurring in, for example, mammalian tissue sections. The mass spectrometer is used to determine the molecular weights of the molecules in the surface layers of the tissue. Molecules desorbed from the sample typically are singly protonated, giving an ion at (M + H)+, where M is the molecular mass. The procedure involves coating the tissue section, or a blotted imprint of the section, with a thin layer of energy-absorbing matrix and then analyzing the sample to produce an ordered array of mass spectra, each containing nominal m/z values typically covering a range of over 50 000 Da. Images...

Molecular Imaging of Biological Samples: Localization of Peptides and Proteins Using MALDI-TOF MS

Automated mass spectrometry imaging with a matrix-assisted laser desorption ionization time-of-flight instrument.

A number of different procedures have been developed for use with matrix-assisted laser desorption/ionization mass spectrometry (MALDI/MS) for the analysis of non-covalent protein-protein complexes. These include use of specific matrix and laser combinations, accumulation of "first shot" spectra, modification of pH and solvent conditions during sample preparation and use of cross-linking agents to attach the monomers covalently to each other in the complex. The results have shown the techniques to be effective with some but not all complexes, although cross-linking is the most successful. The physical and chemical nature of the complex is critical and therefore a diversity of approaches is recommended for such studies.

Determination of protein-protein interactions by matrix-assisted laser desorption/ionization mass spectrometry.

We have developed a technique which utilizes matrix-assisted laser desorption mass spectrometry to study the subunit association of proteins. Aqueous protein samples are treated with a dilute solution of glutaraldehyde, a cross-linking agent which reacts with free amino groups on proteins. This agent effectively traps the multimeric form, preventing it from dissociating in the sample preparation and desorption process. Proteins measured include lysozyme, carbonic anhydrase, apomyoglobin, glucose 6-phosphate dehydrogenase, ovine lutropin, yeast alcohol dehydrogenase, avidin and pyruvate kinase. Dimeric and tetrameric complexes up to 250,000 Da have been measured in this manner.

Assessing the multimeric states of proteins: studies using laser desorption mass spectrometry.

The determination of the mass discrimination effect of matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) over a wide m/z range was made by determining the relative amounts of prepurified protein monomer and dimer species present in reconstituted mixtures. Protein complexes were covalently cross-linked with glutaraldehyde and monomeric and dimeric species were separated by gel filtration chromatography. Individual protein species were recombined in specific ratios and analyzed by both MALDI MS and sodium dodecyl sulfate polyacrylamide gel electrophoresis. The contributions of the peak areas of the doubly charged ions and singlycharged dimeric ions generated during the MALDI MS analyses relative to that of the singly-charged [M + H]+ species were measured. The calculation of mass discrimination for two different MALDI time-of-flight instruments showed the loss of a factor of ∼2.6 in relative peak area as the m/z value doubled over the m/z range 30 000–145 000. An overall correction factor for mass discrimination is given which includes effects associated with sample preparation, sample desorption/ion formation, ion transmission and ion detection.

Terry B. Farmer

Papers

Molecular Imaging of Biological Samples: Localization of Peptides and Proteins Using MALDI-TOF MS

Automated mass spectrometry imaging with a matrix-assisted laser desorption ionization time-of-flight instrument.

Determination of protein-protein interactions by matrix-assisted laser desorption/ionization mass spectrometry.

Assessing the multimeric states of proteins: studies using laser desorption mass spectrometry.

Mass discrimination in matrix‐assisted laser desorption ionization time‐of‐flight mass spectrometry: A study using cross‐linked oligomeric complexes