Imaging with mass spectrometry: Which ionization technique is best?

The use of mass spectrometry (MS) to acquire molecular images of biological tissues and other substrates has developed into an indispensable analytical tool over the past 25 years. Imaging mass spectrometry technologies are widely used today to study the in situ spatial distributions for a variety of analytes. Early MS images were acquired using secondary ion mass spectrometry and matrix-assisted laser desorption/ionization. Researchers have also designed and developed other ionization techniques in recent years to probe surfaces and generate MS images, including desorption electrospray ionization (DESI), nanoDESI, laser ablation electrospray ionization, and infrared matrix-assisted laser desorption electrospray ionization. Investigators now have a plethora of ionization techniques to select from when performing imaging mass spectrometry experiments. This brief perspective will highlight the utility and relative figures of merit of these techniques within the context of their use in imaging mass spectrometry.     

Recent advances of ambient ionization mass spectrometry imaging in clinical research

The structural information and spatial distribution of molecules in biological tissues are closely related to the potential molecular mechanisms of disease origin, transfer, and classification. Ambient ionization mass spectrometry imaging is an effective tool that provides molecular images while describing in situ information of biomolecules in complex samples, in which ionization occurs at atmospheric pressure with the samples being analyzed in the native state. Ambient ionization mass spectrometry imaging can directly analyze tissue samples at a fairly high resolution to obtain molecules in situ information on the tissue surface to identify pathological features associated with a disease, resulting in the wide applications in pharmacy, food science, botanical research, and especially clinical research. Herein, novel ambient ionization techniques, such as techniques based on spray and solid‐liquid extraction, techniques based on plasma desorption, techniques based on laser desorption ablation, and techniques based on acoustic desorption were introduced, and the data processing of ambient ionization mass spectrometry imaging was briefly reviewed. Besides, we also highlight recent applications of this imaging technology in clinical researches and discuss the challenges in this imaging technology and the perspectives on the future of the clinical research.     

Biological tissue imaging with time-of-flight secondary ion mass spectrometry and cluster ion sources

Time-of-flight secondary ion mass spectrometry (TOF-SIMS) using liquid metal ion guns (LMIGs) is now sensitive enough to produce molecular-ion images directly from biological tissue samples. Primary cluster ions strike a spot on the sample to produce a mass spectrum. An image of this sample is achieved by rastering the irradiated point over the sample surface. The use of secondary ion mass spectrometry for mapping biological tissue surfaces provides unique analytical capabilities; in particular, it enables in a single acquisition a large variety of biological compounds to be localised on a micrometer scale and scrutinised for colocalisations. Without any treatment of the sample, this method is fully compatible with subsequent and complementary analyses like fluorescence microscopy, histochemical staining, or even matrix-assisted laser desorption/ionisation imaging. Basic physical concepts, required instrumentation (ion source and mass analyzer), sample preparation methods, image acquisition, image processing, and emerging biological applications will be described and discussed.     

Secondary ion and laser ablation mass spectrometry for the quantitative characterization of styrene-butadiene copolymers

Styrene-butadiene copolymers were analyzed by static secondary ion mass spectrometry (S-SIMS) and laser ablation Fourier transform ion cyclotron resonance mass spectrometry (LA-FTICRMS) to obtain quantitative information based on specific ions. Silver deposition was performed on polystyrene, butadiene rubber and styrene-butadiene rubber. Under these experimental conditions, new secondary ions were detected, in particular silver-cationized butadiene [M(butadiene) - Ag](+) and styrene [M(styrene) - Ag](+) monomers. In contrast, LA-FTICRMS experiments did not require pretreatment. At high laser power density, UV photons (193, 266 and 355 nm) allowed the detection of styrene and butadiene monomers at m/z 104 and 54, respectively. The use of the observed ions by SIMS or LA-FTICRMS ensures that quantitative information on the relative distribution of each monomer is obtained. However, the silver coating thickness in the SIMS experiment seems to have an important influence on the quantitative information obtained. For LA-FTICRMS experiments, the best results are obtained at a wavelength of 355 nm.     

Mass spectrometric analysis of ceramic components for solid oxide fuel cells

For the determination of trace impurities in ceramic components of solid oxide fuel cells (SOFCs), some mass spectrometric methods have been applied such as spark source mass spectrometry (SSMS), laser ionization mass spectrometry (LIMS), laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) and inductively coupled plasma mass spectrometry (ICP-MS). Due to a lack of suitable standard reference materials for quantifying of analytical results on La _x Sr _y MnO₃ cathode material a matrix-matched synthetic standard-high purity initial compounds doped with trace elements-was prepared in order to determine the relative sensitivity coefficients in SSMS and LA-ICP-MS. Radiofrequency glow discharge mass spectrometry (rf-GDMS) was developed for trace analysis and depth profiling of thick non-conducting layers. Surface analytical techniques, such as secondary ion mass spectrometry (SIMS) and sputtered neutral mass spectrometry (SNMS), were used to determine the element distribution on surfaces (homogeneity) and the surface contaminants of SOFC ceramic layers.Dedicated to Professor Dr. rer. nat. Hubertus Nickel on the occasion of his 65th birthday     

Ambient imaging mass spectrometry by electrospray ionization using solid needle as sampling probe

Although being an atmospheric pressure ion source, electrospray ionization (ESI) has rarely been used directly for ambient imaging mass spectrometry because the sample has to be introduced as liquid solution through the capillary. Instead of capillary, probe electrospray ionization (PESI), which has been developed recently, uses a solid needle as the sampling probe, as well as the electrospray emitter, and has been applied not only for liquid solutions but also for the direct sampling on wet samples. Biological tissues are composed of cells that contain 70–90% water, and when the surface is probed by the needle tip, the biological fluid adhering to the needle can be electrosprayed directly or assisted by additional solvent added onto the needle surface. Here, we demonstrate ambient imaging mass spectrometry of mouse brain section using PESI, incorporated with an auxiliary heated capillary sprayer. The solvent vapor generated from the sprayer condensed on the needle tip, re‐dissolving the adhered sample, and at the same time, providing an indirect means for needle cleaning. The histological sections were prepared by fixation using paraformaldehyde, and the spatial analysis was automated by maintaining an equal sampling depth into the sample in addition to raster scan. Phospholipids and galactosylceramides were readily detected from the mouse brain section in the positive ion mode, and were mapped with 60 µm lateral resolution to form mass spectrometric images. Copyright © 2009 John Wiley & Sons, Ltd.     

Atmospheric pressure ionization–tandem mass spectrometry of the phenicol drug family

In this work, the mass spectrometry behaviour of the veterinary drug family of phenicols, including chloramphenicol (CAP) and its related compounds thiamphenicol (TAP), florfenicol (FF) and FF amine (FFA), was studied. Several atmospheric pressure ionization sources, electrospray (ESI), atmospheric pressure chemical ionization and atmospheric pressure photoionization were compared. In all atmospheric pressure ionization sources, CAP, TAP and FF were ionized in both positive and negative modes; while for the metabolite FFA, only positive ionization was possible. In general, in positive mode, [M + H]⁺ dominated the mass spectrum for FFA, while the other compounds, CAP, TAP and FF, with lower proton affinity showed intense adducts with species present in the mobile phase. In negative mode, ESI and atmospheric pressure photoionization showed the deprotonated molecule [M–H]^?, while atmospheric pressure chemical ionization provided the radical molecular ion by electron capture. All these ions were characterized by tandem mass spectrometry using the combined information obtained by multistage mass spectrometry and high‐resolution mass spectrometry in a quadrupole‐Orbitrap instrument. In general, the fragmentation occurred via cyclization and losses or fragmentation of the N‐(alkyl)acetamide group, and common fragmentation pathways were established for this family of compounds. A new chemical structure for the product ion at m/z 257 for CAP, on the basis of the MS³ and MS⁴ spectra is proposed. Thermally assisted ESI and selected reaction monitoring are proposed for the determination of these compounds by ultra high‐performance liquid chromatography coupled to tandem mass spectrometry, achieving instrumental detection limits down to 0.1 pg. Copyright © 2013 John Wiley & Sons, Ltd.     

Infrared Laser Ablation Sample Transfer for MALDI and Electrospray

We have used an infrared laser to ablate materials under ambient conditions that were captured in solvent droplets. The droplets were either deposited on a MALDI target for off-line analysis by MALDI time-of-flight mass spectrometry or flow-injected into a nanoelectrospray source of an ion trap mass spectrometer. An infrared optical parametric oscillator (OPO) laser system at 2.94 μm wavelength and approximately 1 mJ pulse energy was focused onto samples for ablation at atmospheric pressure. The ablated material was captured in a solvent droplet 1–2 mm in diameter that was suspended from a silica capillary a few millimeters above the sample target. Once the sample was transferred to the droplet by ablation, the droplet was deposited on a MALDI target. A saturated matrix solution was added to the deposited sample, or in some cases, the suspended capture droplet contained the matrix. Peptide and protein standards were used to assess the effects of the number of IR laser ablation shots, sample to droplet distance, capture droplet size, droplet solvent, and laser pulse energy. Droplet collected samples were also injected into a nanoelectrospray source of an ion trap mass spectrometer with a 500 nL injection loop. It is estimated that pmol quantities of material were transferred to the droplet with an efficiency of approximately 1%. The direct analysis of biological fluids for off-line MALDI and electrospray was demonstrated with blood, milk, and egg. The implications of this IR ablation sample transfer approach for ambient imaging are discussed.     

一种新型二次离子质谱的一次离子源及其离子光学系统

本研究设计了一种新型用于二次离子质谱的一次离子源及其离子光学系统.通过此一次离子源,大气压下产生的一次离子可以被加速、聚焦并传输到位于真空条件下的样品表面并电离样品得到可供质谱仪分析的二次离子.通过理论模拟结合实验系统研究了此一次离子源的主要组成部分——离子光学系统的原理、结构和性能.以电喷雾电离源为例,成功地将大气压...     

Laser Mass Spectrometry of Organophosphorus Pesticides and Related Compounds

Abstract

Laser mass spectra obtained for 20 organophosphorus (OP) compounds were systematically evaluated for groups containing analogous structural features. Variations in fragmentation can be understood based on simple organic reactions. While detailed mechanistic interpretations of the laser mass spectra (LMS) were not possible, the qualitative features in the LMS obtained from five compounds, not in the original set, could be predicted based on the characteristics of the other OP compounds studied. The success of the prediction lends credence to the qualitative models developed for rationalizing the LMS. A specific feature in the LMS of aromatic thionophosphates is a thiono-thiolo rearrangement. Detailed investigation into the phenomena involved comparison of LMS obtained from aromatic thionophosphates with spectra from electron impact, chemical ionization, field desorption, and secondary ion mass spectrometry. These results led to the conclusion that the rearrangement in laser mass spectrometry must occur during volatilization while the molecule/ion is in the “cloud” present immediately above the laser impact area.     

同位素质谱和无机质谱

本文是《分析试验室》定期评述中“同位素质谱和无机质谱”的第四篇评述，评述的范围是１９９４年１１月至１９９６年１０月我国气体同位素质谱，热电离同位素质谱，加速器质谱，火花源质谱，电感耦合等离子体质谱，辉光放电质谱，同位素稀释质谱，二次离子质谱，激光共振电离子飞行时间质谱，电子探针，质子探针，激光探针和它们在地学，核科学，环境科学，材料学，计理学，医学和生命科学中的应用，引用文献１４９篇。     

Laserspray ionization (LSI) ion mobility spectrometry (IMS) mass spectrometry

A simple device is described for desolvation of highly charged matrix/analyte clusters produced by laser ablation leading to multiply charged ions that are analyzed by ion mobility spectrometry-mass spectrometry. Thus, for example, highly charged ions of ubiquitin and lysozyme are cleanly separated in the gas phase according to size and mass (shape and molecular weight) as well as charge using Tri-Wave ion mobility technology coupled to mass spectrometry. This contribution confirms the mechanistic argument that desolvation is necessary to produce multiply charged matrix-assisted laser desorption/ionization (MALDI) ions and points to how these ions can be routinely formed on any atmospheric pressure mass spectrometer.     

Laserspray ionization using an atmospheric solids analysis probe for sample introduction

A newly introduced high sensitivity laserspray (LSI) mass spectrometry (MS) method that uses laser ablation of a matrix/analyte mixture at atmospheric pressure (AP) to obtain multiply charged ions from nonvolatile as well as high-mass compounds is now implemented using a simple probe device. The probe used in the LSI approach was originally designed for sample introduction into an AP ionization source using the atmospheric solids analysis probe (ASAP) method. Multiply charged mass spectra of peptides and proteins in 2,5-dihydroxybenzoic acid matrix were readily obtained on two mass spectrometers from different manufacturers with sample introduction using melting point tubes. Here we demonstrate rapid analysis by placing four peptide and protein samples on a single melting point tube. Mass spectra were obtained at high-resolution and using ion mobility spectrometry/MS.     

Laser desorption-atmospheric pressure chemical ionization: a novel ion source for the direct coupling of polyacrylamide gel electrophoresis to mass spectrometry

Laser desorption-atmospheric pressure chemical ionization-mass spectrometry (LD-APCI-MS) is presented for the atmospheric pressure (AP) sampling of tryptic peptides directly from a polyacrylamide gel. In contrast to other gel sampling mass spectrometric approaches, this technique does not require the addition of any exogenous matrices to the gel to assist with ionization. In this arrangement, a CO(2) laser at 10.6 micro m is used to desorb intact neutral peptide molecules from the gel, followed by ionization in the gas-phase with APCI. The ions are then sampled via a heated capillary inlet and transferred to a quadrupole ion trap mass spectrometer for mass analysis. Preliminary results suggest the polyacrylamide gel electrophoresis-LD-APCI-MS technique provides several advantages that could translate into a more convenient, robust methodology for the rapid identification and characterization of proteins. Finally, strategies regarding the further development of the method are presented.     

Profiling and imaging of tissues by imaging ion mobility-mass spectrometry

Molecular profiling and imaging mass spectrometry (IMS) of tissues can often result in complex spectra that are difficult to interpret without additional information about specific signals. This report describes increasing data dimensionality in IMS by combining two-dimensional separations at each spatial location on the basis of imaging ion mobility-mass spectrometry (IM-MS). Analyte ions are separated on the basis of both ion-neutral collision cross section and m/z, which provides rapid separation of isobaric, but structurally distinct ions. The advantages of imaging using ion mobility prior to MS analysis are demonstrated for profiling of human glioma and selective lipid imaging from rat brain.     

Direct imaging of single cells and tissue at sub‐cellular spatial resolution using transmission geometry MALDI MS

The need of cellular and sub‐cellular spatial resolution in laser desorption ionization (LDI)/matrix‐assisted LDI (MALDI) imaging mass spectrometry (IMS) necessitates micron and sub‐micron laser spot sizes at biologically relevant sensitivities, introducing significant challenges for MS technology. To this end, we have developed a transmission geometry vacuum ion source that allows the laser beam to irradiate the back side of the sample. This arrangement obviates the mechanical/ion optic complications in the source by completely separating the optical lens and ion optic structures. We have experimentally demonstrated the viability of transmission geometry MALDI MS for imaging biological tissues and cells with sub‐cellular spatial resolution. Furthermore, we demonstrate that in conjunction with new sample preparation protocols, the sensitivity of this instrument is sufficient to obtain molecular images at sub‐micron spatial resolution. Copyright © 2012 John Wiley & Sons, Ltd.     

Nuclear Magnetic Resonance Structure Elucidation of Peptide b2 Ions

Tandem mass spectrometry (MS/MS) is powerful for chemical identification but it is still insufficient for explicit ion structure determination. A strategy is introduced to elucidate MS fragment ion structures using NMR spectroscopy for the first time. In our experiments, precursor ions are dissociated at atmospheric pressure and the resulting fragment ions are identified by mass spectrometry but collected outside the mass spectrometer, making the subsequent NMR measurements possible. This new strategy has been applied to determine the chemical structure of the characteristic b₂ fragment ion, a subject of longstanding debate in MS‐based proteomics.     

Mass spectrometry based cellular phosphoinositides profiling and phospholipid analysis: A brief review

Phospholipids are key components of cellular membrane and signaling. Among cellular phospholipids, phosphoinositides, phosphorylated derivatives of phosphatidylinositol are important as a participant in essential metabolic processes in animals. However, due to its low abundance in cells and tissues, it is difficult to identify the composition of phosphoinositides. Recent advances in mass spectrometric techniques, combined with established separation methods, have allowed the rapid and sensitive detection and quantification of a variety of lipid species including phosphoinositides. In this mini review, we briefly introduce progress in profiling of cellular phosphoinositides using mass spectrometry. We also summarize current progress of matrices development for the analysis of cellular phospholipids using matrix-assisted laser desorption/ionization mass spectrometry. The phosphoinositides profiling and phospholipids imaging will help us to understand how they function in a biological system and will provide a powerful tool for elucidating the mechanism of diseases such as diabetes, cancer and neurodegenerative diseases. The investigation of cellular phospholipids including phosphoinositides using electrospray ionization mass spectrometry and matrix-assisted laser desorption/ionization mass spectrometry will suggest new insights on human diseases, and on clinical application through drug development of lipid related diseases.     

Determination of polar organic compounds in atmospheric aerosols by gas chromatography with ion trap tandem mass spectrometry

A gas chromatography with ion trap mass spectrometry method has been developed and validated for the analysis of 27 polar organic compounds in atmospheric aerosols. The target analytes were low‐molecular‐weight carboxylic acids and methoxyphenols, as relevant markers of source emissions and photochemical processes of organic aerosols. The operative parameters were optimized in order to achieve the best sensitivity and selectivity for the analysis. In comparison with the previous gas chromatography with mass spectrometry procedure based on single ion monitoring detection, the tandem mass spectrometry technique increased the analytical sensitivity by reducing detection limits for standard solutions from 1–2.6 to 0.1–0.4 ng/μL ranges (concentrations in the injected solution). In addition, it enhanced selectivity by reducing matrix interferences and chemical noise in the chromatogram. The applicability of the developed method in air quality monitoring campaigns was effectively checked by analyzing environmental samples collected in the Po Valley (Northern Italy) in different seasons. The obtained results indicate that the ion trap mass spectrometer may be an ideal alternative to high‐resolution mass spectrometers for the user‐friendly and cost‐effective determination of a wide range of molecular tracers in airborne particulate matter.     

Instrument design and characterization for high resolution MALDI-MS imaging of tissue sections

In previous work, we have reported using a MALDI imaging time-of-flight mass spectrometer for the detection of protein ions from tissue sections with spatial resolution of 25 microm. We present here imaging mass spectrometry results obtained with a high-resolution scanning MALDI time-of-flight mass spectrometer, equipped with a coaxial laser illumination ion source, capable of achieving irradiation areas as small as 40 microm(2) (ca 7 microm diameter). MALDI-generated analyte ion signals from these very small irradiation volumes can be observed in a molecular weight range up to 27,000. High-resolution imaging mass spectrometry images were successfully generated from matrix thin film samples and tissue sections with scanning resolutions at and below 10 microm. This work also provides fundamental characterization of the ion signal dependence as a function of various focus and fluence parameters that will be required for extension to tissue imaging at the subcellular level.