Compression strategies for the chemometric analysis of mass spectrometry imaging data

Application of chemometric methods to mass spectrometry imaging (MSI) data faces a bottleneck concerning the vast size of the experimental data sets. This drawback is critical when considering high‐resolution mass spectrometry data, which provide several thousand points for each considered pixel. In this work, different approaches have been tested to reduce the size of the analyzed data with the aim to allow the subsequent application of typical chemometric methods for image analysis. The standard approach for MSI data compression consists in binning mass spectra for each pixel to reduce the number of m/z values. In this work, a method is proposed to handle the huge size of MSI data based on the adaptation of a liquid chromatography‐mass spectrometry data compression method by the detection of regions of interest. Results showed that both approaches achieved high compression rates, although the proposed regions of interest–based method attains this reduction requiring lower computational requirements and keeping utter spectral information. For instance, typical compression rate reached values higher than 90% without loss of information in images and spectra.     

Novel matrix strategies for improved ionization and spatial resolution using IR-MALDESI mass spectrometry imaging

In mass spectrometry imaging (MSI) applications of infrared matrix-assisted laser desorption electrospray ionization (IR-MALDESI), an exogenous ice layer is the gold standard for an energy-absorbing matrix. However, the formation of the ice matrix requires additional time and instrument hardware, so glycerol was investigated herein as an alternative to the ice matrix to potentially improve spatial resolution and ionization, while decreasing experiment time. Glycerol solutions of varying concentrations were sprayed over top of rat liver tissue sections for analysis by IR-MALDESI and compared to the typical ice matrix condition. Additionally, we tested if combining the ice matrix and glycerol matrix would further improve analyses. Matrix conditions were evaluated by comparing ion abundance of six lipid species, the laser ablation spot diameter, and number of METASPACE annotations. The ion abundances were also normalized to the volume of tissue ablated to correct for lower abundance values due to less ablated tissue. It was observed that utilizing a 50% glycerol matrix without ice provides improved spatial resolution with lipid abundances and annotations comparable to the ice matrix standard, while decreasing the time required to complete an IR-MALDESI tissue imaging experiment.     

Development of quantitative imaging mass spectrometry (q-IMS) for drug visualization using animal tissues

Quantitative imaging mass spectrometry (q-IMS) is a frontier topic of research in drug analysis. Although many q-IMS methodologies have been reported, validation of the method is insufficient. We have investigated the feasibility of coupling q-IMS with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to develop a verifiable method. The approach combines quantitative LC-MS/MS information with the spatial distribution information obtained by IMS. This paper compares measured drug quantities with those estimated using IMS. The target drug, erlotinib, is a tyrosine kinase inhibitor of non-small-cell lung cancer. The quantitative accuracy of our q-IMS method in an animal model study is approximately 17%. Measurements were conducted on mouse liver and brain tissues before and after erlotinib administration. Erlotinib is delivered to the brain, although the concentration is 10⁴ times smaller than that found in the liver.     

Multiplex mass spectrometry imaging for latent fingerprints

We have previously developed in‐parallel data acquisition of orbitrap mass spectrometry (MS) and ion trap MS and/or MS/MS scans for matrix‐assisted laser desorption/ionization MS imaging (MSI) to obtain rich chemical information in less data acquisition time. In the present study, we demonstrate a novel application of this multiplex MSI methodology for latent fingerprints. In a single imaging experiment, we could obtain chemical images of various endogenous and exogenous compounds, along with simultaneous MS/MS images of a few selected compounds. This work confirms the usefulness of multiplex MSI to explore chemical markers when the sample specimen is very limited. Copyright © 2013 John Wiley & Sons, Ltd.     

Design and performance of air flow-assisted ionization imaging mass spectrometry system

The imaging mass spectrometry（IMS） technology has experienced a rapid development in recent years.A new IMS technology which is based on air flow assisted ionization（AFAI） was reported.It allows for the convenient pretreatment of the samples and can image a large area of sample in a single measurement with high sensitivity.The AFAI in DESI mode was used as the ion source in this paper.The new IMS method is named AFADESI-IMS.The adoption of assisted air flow makes the sample pretreatment easy and convenient.An optimization of the distance between the ion transport tube and MS orifice increases the sensitivity of the system.For data processing,a program based on MATLAB with the function of numerical analysis was developed.A theoretical imaging resolution of a few hundred microns can be achieved.The composite AFAI-IMS images of different target analytes were imaged with high sensitivity.A typical AFAI-IMS image of the whole-body section of a rat was obtained in a single analytical measurement.The ability to image a large area for relevant samples in a single measurement with high sensitivity and repeatability is a significant advantage.The method has enormous potentials in the MS imaging of large and complicated samples.     

Time of acquisition and high spatial resolution mass spectrometry imaging

The field of mass spectrometry imaging (MSI) is constantly evolving to analyze a diverse array of biological systems. A common goal is the need to resolve cellular and subcellular heterogeneity with high spatial resolution. As the field continues to progress towards high spatial resolution, other parameters must be considered when developing a practical method. Here, we discuss the impacts of high spatial resolution on the time of acquisition and the associated implications they have on an MSI analysis (e.g., area of the region of interest). This work presents a brief tutorial serving to evaluate high spatial resolution MSI relative to time of acquisition and data file size.     

质谱成像分析技术、方法与应用进展

质谱成像技术作为分子成像及质谱领域的研究前沿和热点,近几年受到高度关注并得到迅速发展.本文针对质谱成像技术、方法及其应用的新进展进行了综述与展望,介绍了中国学者近几年在质谱成像分析新技术及其应用方面取得的重要进展.     

Myofiber metabolic type determination by mass spectrometry imaging

Matrix assisted laser desorption/ionization (MALDI) mass spectrometry imaging is a powerful tool that opens new research opportunities in the field of biology. In this work, predictive model was developed to discriminate metabolic myofiber types using the MALDI spectral data. Rat skeletal muscles are constituted of type I and type IIA fiber, which have an oxidative metabolism for glycogen degradation, and type IIX and type IIB fiber which have a glycolytic metabolism, present in different proportions according to the muscle function and physiological state. So far, myofiber type is determined by histological methods that are time consuming. Thanks to the predictive model, we were able to predict not only the metabolic fiber type but also their location, on the same muscle section that was used for MALDI imaging. Copyright © 2017 John Wiley & Sons, Ltd.     

Fabrication of homogenous three‐dimensional biomimetic tissue for mass spectrometry imaging

Reference samples are essential for mass spectrometric method optimization, data quality control, and target analyte quantitation. However, it is highly challenging to prepare an ideal homogeneous, standard‐spiked tissue sample for mass spectrometry imaging (MSI) research. Herein, we present a standard‐spiked 3D biomimetic tissue model fabricated with native cells, homogenate matrix, and biocompatible polymer. Unlike traditional homogenized tissue surrogates or those constructed with “on‐tissue” or “under‐tissue” micropipetting strategies, this simulated tissue shares both structural integrity of cells and homogeneous properties of matrix. As a result, analyte standards could undergo more in‐depth incorporation and has a more comparable native status with a real tissue. Series of tissue sections made from the 3D tissue model were proven to be feasible and useful for the parameter optimization, analyte quantitation, and calibration curve fitting for the air‐flow assisted desorption electrospray ionization MSI. Additionally, by analyzing the quality control model sections, we proposed a median principal component score calibration and demonstrated that this method can normalize instrumental fluctuations to stable levels in a large‐scale untargeted MSI experiments for the reliable metabolomic biomarker discovery. Thus, these results indicated that the standard‐spiked 3D biomimetic tissue has convincing significance in MSI analysis     

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.     

Uncovering matrix effects on lipid analyses in MALDI imaging mass spectrometry experiments

The specific matrix used in matrix‐assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) can have an effect on the molecules ionized from a tissue sample. The sensitivity for distinct classes of biomolecules can vary when employing different MALDI matrices. Here, we compare the intensities of various lipid subclasses measured by Fourier transform ion cyclotron resonance (FT‐ICR) IMS of murine liver tissue when using 9‐aminoacridine (9AA), 5‐chloro‐2‐mercaptobenzothiazole (CMBT), 1,5‐diaminonaphthalene (DAN), 2,5‐Dihydroxyacetophenone (DHA), and 2,5‐dihydroxybenzoic acid (DHB). Principal component analysis and receiver operating characteristic curve analysis revealed significant matrix effects on the relative signal intensities observed for different lipid subclasses and adducts. Comparison of spectral profiles and quantitative assessment of the number and intensity of species from each lipid subclass showed that each matrix produces unique lipid signals. In positive ion mode, matrix application methods played a role in the MALDI analysis for different cationic species. Comparisons of different methods for the application of DHA showed a significant increase in the intensity of sodiated and potassiated analytes when using an aerosol sprayer. In negative ion mode, lipid profiles generated using DAN were significantly different than all other matrices tested. This difference was found to be driven by modification of phosphatidylcholines during ionization that enables them to be detected in negative ion mode. These modified phosphatidylcholines are isomeric with common phosphatidylethanolamines confounding MALDI IMS analysis when using DAN. These results show an experimental basis of MALDI analyses when analyzing lipids from tissue and allow for more informed selection of MALDI matrices when performing lipid IMS experiments.     

SMART: A data reporting standard for mass spectrometry imaging By Ying Xi,Alexandria L. Sohn,Alena N. Joignant,Stephanie M. Cologna,Boone M. Prentice and David C. Muddiman

Mass spectrometry imaging (MSI) is an important analytical technique that simultaneously reports the spatial location and abundance of detected ions in biological, chemical, clinical, and pharmaceutical studies. As MSI grows in popularity, it has become evident that data reporting varies among different research groups and between techniques. The lack of consistency in data reporting inherently creates additional challenges in comparing intra- and inter-laboratory MSI data. In this tutorial, we propose a unified data reporting system, SMART, based on the common features shared between techniques. While there are limitations to any reporting system, SMART was decided upon after significant discussion to more easily understand and benchmark MSI data. SMART is not intended to be comprehensive but rather capture essential baseline information for a given MSI study; this could be within a study (e.g., effect of spot size on the measured ion signals) or between two studies (e.g., different MSI platform technologies applied to the same tissue type). This tutorial does not attempt to address the confidence with which annotations are made nor does it deny the importance of other parameters that are not included in the current SMART format. Ultimately, the goal of this tutorial is to discuss the necessity of establishing a uniform reporting system to communicate MSI data in publications and presentations in a simple format to readily interpret the parameters and baseline outcomes of the data.     

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.     

MASSyPup — an ‘Out of the Box’ solution for the analysis of mass spectrometry data

Mass spectrometry has evolved to a key technology in the areas of metabolomics and proteomics. Centralized facilities generate vast amount of data, which frequently need to be processed off‐site. Therefore, the distribution of data and software, as well as the training of personnel in the analysis of mass spectrometry data, becomes increasingly important. Thus, we created a comprehensive collection of mass spectrometry software which can be run directly from different media such as DVD or USB without local installation. MASSyPup is based on a Linux Live distribution and was complemented with programs for conversion, visualization and analysis of mass spectrometry (MS) data. A special emphasis was put on protein analysis and proteomics, encompassing the measurement of complete proteins, the identification of proteins based on Peptide Mass Fingerprints (PMF) or LC‐MS/MS data, and de novo sequencing. Another focus was directed to the study of metabolites and metabolomics, covering the detection, identification and quantification of compounds, as well as subsequent statistical analyses. Additionally, we added software for Mass Spectrometry Imaging (MSI), including hardware support for self‐made MSI devices. MASSyPup represents a ‘ready to work’ system for teaching or MS data analysis, but also represents an ideal platform for the distribution of MS data and the development of related software. The current Live DVD version can be downloaded free of charge from http://www.bioprocess.org/massypup . Copyright © 2014 John Wiley & Sons, Ltd.     

Decellularization of intact tissue enables MALDI imaging mass spectrometry analysis of the extracellular matrix

Matrix‐assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is a powerful molecular mapping technology that offers unbiased visualization of the spatial arrangement of biomolecules in tissue. Although there has been a significant increase in the number of applications employing this technology, the extracellular matrix (ECM) has received little attention, likely because ECM proteins are mostly large, insoluble and heavily cross‐linked. We have developed a new sample preparation approach to enable MALDI IMS analysis of ECM proteins in tissue. Prior to freezing and sectioning, intact tissues are decellularized by incubation in sodium dodecyl sulfate. Decellularization removes the highly abundant, soluble species that dominate a MALDI IMS spectrum while preserving the structural integrity of the ECM. In situ tryptic hydrolysis and imaging of tryptic peptides are then carried out to accommodate the large sizes of ECM proteins. This new approach allows the use of MALDI IMS for identification of spatially specific changes in ECM protein expression and modification in tissue. Copyright © 2015 John Wiley & Sons, Ltd.     

介质阻挡放电离子源质谱分析

敞开式离子化质谱可对表面样品进行直接快速分析而受到关注,成为质谱分析的热点研究方向.介质阻挡放电离子源是一种基于等离子体放电机理的敞开式离子源,近年来得到了较快的发展.本文着重介绍该离子源的基本原理、性能特征以及应用进展,并对其发展趋势进行了展望.     

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.     

Mapping the fly Malpighian tubule lipidome by imaging mass spectrometry

Matrix‐assisted laser/desorption ionization imaging mass spectrometry (MALDI IMS) is an analytical technique for understanding the spatial distribution of biomolecules across a sample surface. Originally employed for mammalian tissues, this technology has been adapted to study specimens as diverse as microbes and cell cultures, food such as strawberries, and invertebrates including the vinegar fly Drosophila melanogaster. As an ideal model organism, Drosophila has brought greater understanding about conserved biological processes, organism development, and diseased states and even informed management practices of agriculturally and environmentally important species. Drosophila displays anatomically separated renal (Malpighian) tubules that are the physiological equivalent to the vertebrate nephron. Insect Malpighian tubules are also responsible for pesticide detoxification. In this article, we first describe an effective workflow and sample preparation method to study the phospholipid distribution of the Malpighian tubules that initially involves the manual microdissection of the tubules in saline buffer followed by a series of washes to remove excess salt and enhances the phospholipid signals prior to matrix deposition and IMS at 25‐μm spatial resolution. We also established a complementary methodology for lipid IMS analysis of whole‐body fly sections using a dual‐polarity data acquisition approach at the same spatial resolution after matrix deposition by sublimation. Both procedures yield rich signal profiles from the major phospholipid classes. The reproducibility and high‐quality results offered by these methodologies enable cohort studies of Drosophila through MALDI IMS.     

The evolving field of imaging mass spectrometry and its impact on future biological research

Within the past decade, imaging mass spectrometry (IMS) has been increasingly recognized as an indispensable technique for studying biological systems. Its rapid evolution has resulted in an impressive array of instrument variations and sample applications, yet the tools and data are largely confined to specialists. It is therefore important that at this junction the IMS community begin to establish IMS as a permanent fixture in life science research thereby making the technology and/or the data approachable by non-mass spectrometrists, leading to further integration into biological and clinical research. In this perspective article, we provide insight into the evolution and current state of IMS and propose some of the directions that IMS could develop in order to stay on course to become one of the most promising new tools in life science research.     

Monitoring time‐dependent degradation of phospholipids in sectioned tissues by MALDI imaging mass spectrometry

Imaging mass spectrometry (IMS) is useful for visualizing the localization of phospholipids on biological tissue surfaces creating great opportunities for IMS in lipidomic investigations. With advancements in IMS of lipids, there is a demand for large‐scale tissue studies necessitating stable, efficient and well‐defined sample handling procedures. Our work within this article shows the effects of different storage conditions on the phospholipid composition of sectioned tissues from mouse organs. We have taken serial sections from mouse brain, kidney and liver thaw mounted unto ITO‐coated glass slides and stored them under various conditions later analyzing them at fixed time points. A global decrease in phospholipid signal intensity is shown to occur and to be a function of time and temperature. Contrary to the global decrease, oxidized phospholipid and lysophospholipid species are found to increase within 2 h and 24 h, respectively, when mounted sections are kept at ambient room conditions. Imaging experiments reveal that degradation products increase globally across the tissue. Degradation is shown to be inhibited by cold temperatures, with sample integrity maintained up to a week after storage in ?80 °C freezer under N₂ atmosphere. Overall, the results demonstrate a timeline of the effects of lipid degradation specific to sectioned tissues and provide several lipid species which can serve as markers of degradation. Importantly, the timeline demonstrates oxidative sample degradation begins appearing within the normal timescale of IMS sample preparation of lipids (i.e. 1–2 h) and that long‐term degradation is global. Taken together, these results strengthen the notion that standardized procedures are required for phospholipid IMS of large sample sets, or in studies where many serial sections are prepared together but analyzed over time such as in 3‐D IMS reconstruction experiments. Copyright © 2014 John Wiley & Sons, Ltd.