Ambient Ionization Mass Spectrometry Imaging Research Articles

Hardware and software solutions for implementing nanospray desorption electrospray ionization (nano-DESI) sources on commercial mass spectrometers.

Nanospray desorption electrospray ionization (nano-DESI) is an ambient ionization mass spectrometry imaging (MSI) approach that enables spatial mapping ofbiological and environmental samples with high spatial resolution and throughput. Because nano-DESI has not yet been commercialized, researchers develop their own sources and interface them with different commercial mass spectrometers. Previously, several protocols focusing on the fabrication of nano-DESI probes have been reported. In this tutorial, we discuss different hardware requirements for coupling the nano-DESI source to commercial mass spectrometers, such as the safety interlock, inlet extension, and contact closure. In addition, we describe the structure of our custom software for controlling the nano-DESI MSI platform and provide detailed instructions for its usage. With this tutorial, interested researchers should be able to implement nano-DESI experiments in their labs.

Contrast optimization of mass spectrometry imaging (MSI) data visualization by threshold intensity quantization (TrIQ).

Mass spectrometry imaging (MSI) enables the unbiased characterization of surfaces with respect to their chemical composition. In biological MSI, zones with differential mass profiles hint towards localized physiological processes, such as the tissue-specific accumulation of secondary metabolites, or diseases, such as cancer. Thus, the efficient discovery of ‘regions of interest’ (ROI) is of utmost importance in MSI. However, often the discovery of ROIs is hampered by high background noise and artifact signals. Especially in ambient ionization MSI, unmasking biologically relevant information from crude data sets is challenging. Therefore, we implemented a Threshold Intensity Quantization (TrIQ) algorithm for augmenting the contrast in MSI data visualizations. The simple algorithm reduces the impact of extreme values (‘outliers’) and rescales the dynamic range of mass signals. We provide an R script for post-processing MSI data in the imzML community format (https://bitbucket.org/lababi/msi.r) and implemented the TrIQ in our open-source imaging software RmsiGUI (https://bitbucket.org/lababi/rmsigui/). Applying these programs to different biological MSI data sets demonstrated the universal applicability of TrIQ for improving the contrast in the MSI data visualization. We show that TrIQ improves a subsequent detection of ROIs by sectioning. In addition, the adjustment of the dynamic signal intensity range makes MSI data sets comparable.

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.

Precision biomarker discovery powered by microscopy image fusion-assisted high spatial resolution ambient ionization mass spectrometry imaging

Mass spectrometry imaging (MSI) using the ambient ionization technique enables a direct chemical investigation of biological samples with minimal sample pretreatment. However, detailed morphological information of the sample is often lost due to its limited spatial resolution. In this study, predictive high-resolution molecular imaging was produced by the fusion of ambient ionization MSI with optical microscopy of routine hematoxylin and eosin (H&E) staining. Specifically, desorption electrospray ionization (DESI) and nanospray desorption electrospray ionization (nanoDESI) mass spectrometry were employed to visualize lipid and protein species on mice tissue sections. The resulting molecular distributions obtained by ambient ionization MSI-microscopy fusion were verified with matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) MSI and immunohistochemistry (IHC) staining. Label-free molecular imaging with 5-μm spatial resolution can be acquired using DESI and nanoDESI, whereas the typical spatial resolution of ambient ionization MSI was ∼100 μm. In this regard, sharpened molecular histology of tissue sections was achieved, providing complementary references to the pathology. Such a multi-modal integration enables the discovery of potential tumor biomarkers. After image fusion, more than a dozen potential biomarkers on a metastatic mouse lung tissue section and Luminal B breast tumor tissue section were identified.

Ambient ionization mass spectrometry imaging for disease diagnosis: Excitements and challenges

Tissue analysis in histology is extremely important and also considered to be a gold standard to diagnose and prognosticate several diseases including cancer. Intraoperative evaluation of surgical margin of tumor also relies on frozen section histopathology, which is time consuming, challenging and often subjective. Recent development in the ambient ionization mass spectrometry imaging (MSI) technique has enabled us to simultaneously visualize hundreds to thousands of molecules (ion images) in the biopsy specimen, which are strikingly different and more powerful than the single optical tissue image analysis in conventional histopathology. This paper will highlight the emergence of the desorption electrospray ionization MSI (DESI-MSI) technique, which is label-free, requires minimal or no sample preparation and operates under ambient conditions. DESI-MSI can record ion images of lipid/metabolite distributions on biopsy specimens, providing a wealth of diagnostic information based on differential distributions of these molecular species in healthy and unhealthy tissues. Remarkable success of this technology in rapidly evaluating the cancer margin intraoperatively with very high accuracy also promises to bring this imaging technique from bench to bedside.

Desorption Electrospray Ionization Mass Spectrometry Imaging of Proteins Directly from Biological Tissue Sections.

Analysis of large biomolecules including proteins has proven challenging using ambient ionization mass spectrometry imaging techniques. Here, we have successfully optimized desorption electrospray ionization mass spectrometry (DESI-MS) to detect intact proteins directly from tissue sections and further integrated DESI-MS to a high field asymmetric waveform ion mobility (FAIMS) device for protein imaging. Optimized DESI-FAIMS-MS parameters were used to image mouse kidney, mouse brain, and human ovarian and breast tissue samples, allowing detection of 11, 16, 14, and 16 proteoforms, respectively. Identification of protein species detected by DESI-MS was performed on-tissue by top-down ultraviolet photodissociation (UVPD) and collision induced dissociation (CID) as well as using tissue extracts by bottom-up CID and top-down UVPD. Our results demonstrate that DESI-MS imaging is suitable for the analysis of the distribution of proteins within biological tissue sections.

Early Detection of Biofouling on Water Purification Membranes by Ambient Ionization Mass Spectrometry Imaging.

By direct analysis of water purification membranes using ambient ionization mass spectrometry, an attempt has been made to understand the molecular signatures of bacterial fouling. Membrane based purification methods are used extensively in water treatment, and a major challenge for them is biofouling. The buildup of microbes and their extracellular polymeric matrix clog the purification membranes and reduce their efficiency. To understand the early stages of bacterial fouling on water purification membranes, we have used desorption electrospray ionization mass spectrometry (DESI MS), where ion formation occurs in ambient conditions and the ionization event is surface sensitive. Biosurfactants at the air-water interface generated by microorganisms as a result of quorum sensing, influence the water-membrane interface and are important for the bacterial attachment. We show that these biosurfactants produced by bacteria can be indicator molecular species signifying initiation of biofilms on membrane surfaces, demonstrated by specific DESI MS signatures. In Pseudomonas aeruginosa, one of the best studied models for biofilm formation, this process is mediated by rhamnolipids forewarning bacterial fouling. Species dependent variation of such molecules can be used for the precise identification of the microorganisms, as revealed by studies on P. aeroginosa (ATCC 25619). The production of biosurfactants is tightly regulated at the transcriptional level by the quorum-sensing (QS) response. Thus, secretion of these extracellular molecules across the membrane surface allows rapid screening of the biofilm community. We show that, the ambient ionization mass spectrometry can detect certain toxic heavy metals present in water, using surfactant-metal complexes as analytes. We believe that such studies conducted on membranes in various input water streams will help design suitable membrane processes specific to the input streams.

Metabolic Markers and Statistical Prediction of Serous Ovarian Cancer Aggressiveness by Ambient Ionization Mass Spectrometry Imaging.

Ovarian high-grade serous carcinoma (HGSC) results in the highest mortality among gynecological cancers, developing rapidly and aggressively. Dissimilarly, serous borderline ovarian tumors (BOT) can progress into low-grade serous carcinomas and have relatively indolent clinical behavior. The underlying biological differences between HGSC and BOT call for accurate diagnostic methodologies and tailored treatment options, and identification of molecular markers of aggressiveness could provide valuable biochemical insights and improve disease management. Here, we used desorption electrospray ionization (DESI) mass spectrometry (MS) to image and chemically characterize the metabolic profiles of HGSC, BOT, and normal ovarian tissue samples. DESI-MS imaging enabled clear visualization of fine papillary branches in serous BOT and allowed for characterization of spatial features of tumor heterogeneity such as adjacent necrosis and stroma in HGSC. Predictive markers of cancer aggressiveness were identified, including various free fatty acids, metabolites, and complex lipids such as ceramides, glycerophosphoglycerols, cardiolipins, and glycerophosphocholines. Classification models built from a total of 89,826 individual pixels, acquired in positive and negative ion modes from 78 different tissue samples, enabled diagnosis and prediction of HGSC and all tumor samples in comparison with normal tissues, with overall agreements of 96.4% and 96.2%, respectively. HGSC and BOT discrimination was achieved with an overall accuracy of 93.0%. Interestingly, our classification model allowed identification of three BOT samples presenting unusual histologic features that could be associated with the development of low-grade carcinomas. Our results suggest DESI-MS as a powerful approach for rapid serous ovarian cancer diagnosis based on altered metabolic signatures. Cancer Res; 77(11); 2903-13. ©2017 AACR.

Ambient Ionization and FAIMS Mass Spectrometry for Enhanced Imaging of Multiply Charged Molecular Ions in Biological Tissues.

Ambient ionization mass spectrometry imaging (MSI) has been increasingly used to investigate the molecular distribution of biological tissue samples. Here, we report the integration and optimization of desorption electrospray ionization (DESI) and liquid-microjunction surface sampling probe (LMJ-SSP) with a chip-based high-field asymmetric waveform ion mobility spectrometry (FAIMS) device to image metabolites, lipids, and proteins in biological tissue samples. Optimized FAIMS parameters for specific molecular classes enabled semitargeted detection of multiply charged molecular species at enhanced signal-to-noise ratios (S/N), improved visualization of spatial distributions, and, most importantly, allowed detection of species which were unseen by ambient ionization MSI alone. Under static DESI-FAIMS conditions selected for transmission of doubly charged cardiolipins (CL), for example, detection of 71 different CL species was achieved in rat brain, 23 of which were not observed by DESI alone. Diagnostic CL were imaged in a human thyroid tumor sample with reduced interference of isobaric species. LMJ-SSP-FAIMS enabled detection of 84 multiply charged protein ions in rat brain tissue, 66 of which were exclusive to this approach. Spatial visualization of proteins in substructures of rat brain, and in human ovarian cancerous, necrotic, and normal tissues was achieved. Our results indicate that integration of FAIMS with ambient ionization MS allows improved detection and imaging of selected molecular species. We show that this methodology is valuable in biomedical applications of MSI for detection of multiply charged lipids and proteins from biological tissues.

Electrostatic Spray Ionization Mass Spectrometry Imaging

Imaging samples on a surface by mass spectrometry (MS) requires the combination of MS detection with a scanning mode that enables localized desorption and ionization and/or detection of sample analytes with good spatial resolution. We have developed a new mass spectrometry imaging (MSI) method based on electrostatic spray ionization. It works under ambient conditions and can be applied to a wide range of molecules providing quantitative MS analysis even in the presence of salts in excess. 2D MS images of protein and peptide spots, inkjet-printed black dye patterns, and cells were obtained. The presented novel ambient ionization mass spectrometry imaging method can find many applications in analytical and bioanalytical chemistry.

Secondary Metabolites in Fleshy Fruits: Are Adaptive Explanations Needed?

1. Department of Botany, Stockholm University, S-106 91 birds, their presence has been considered to be an evoluStockholm, Sweden; tionary paradox. 2. Department of Ecological Botany, Uppsala University, Cipollini and Levey (1997) provide a list of ingenious Villavagen 14, S-752 36 Uppsala, Sweden and interesting hypotheses focusing on the potential functions of the secondary metabolites for enhancing Submitted December 8, 1997; Accepted April 9, 1998 plant seed dispersal, for example, by providing foraging cues, as regulators of seed germination, as mechanisms to force frugivores not to stay too long at the same plant, as mechanisms to manipulate seeds’ gut passage rate, and as