ORCID Profile
0000-0002-3326-5991
Current Organisation
University of Wollongong
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Analytical Chemistry | Analytical Spectrometry | Medical Biochemistry: Lipids | Characterisation of Biological Macromolecules | Instrumental Methods (excl. Immunological and Bioassay Methods) | Analytical Biochemistry | Nanochemistry and Supramolecular Chemistry
Expanding Knowledge in the Chemical Sciences | Expanding Knowledge in the Biological Sciences | Expanding Knowledge in the Medical and Health Sciences |
Publisher: Proceedings of the National Academy of Sciences
Date: 06-11-2017
Abstract: This work used mass spectrometry imaging to visualize a key bacterial virulence factor, lipid A within tissues infected by Francisella novicida , a model organism for Francisella tularensis ssp. tularensis , the causative agent of tularemia. Lipid components of the host were mapped using the same technique, leading to identification of a lethal role for host lipid metabolism in Francisella infection. Combined with profiling of lipid inflammatory mediators, this work defined the dramatic polarity of the cyclooxygenase-2 axis in this infection and overproduction of the proinflammatory product prostaglandin E2. The host–pathogen lipid interface was reconstructed to demonstrate the spatial organization of the host lipid response in the spleen.
Publisher: American Chemical Society (ACS)
Date: 13-01-2021
Publisher: Elsevier BV
Date: 2020
Publisher: Wiley
Date: 22-12-2020
Publisher: Springer Science and Business Media LLC
Date: 26-12-2019
DOI: 10.1007/S00216-019-02290-3
Abstract: Matrix-assisted laser desorption/ionisation-mass spectrometry imaging (MALDI-MSI) is a powerful technique for visualising the spatial locations of lipids in biological tissues. However, a major challenge in interpreting the biological significance of local lipid compositions and distributions detected using MALDI-MSI is the difficulty in associating spectra with cellular lipid metabolism within the tissue. By-and-large this is due to the typically limited spatial resolution of MALDI-MSI (30–100 μm) meaning in idual spectra represent the average spectrum acquired from multiple adjacent cells, each potentially possessing a unique lipid composition and biological function. The use of overs ling is one promising approach to decrease the s ling area and improve the spatial resolution in MALDI-MSI, but it can suffer from a dramatically decreased sensitivity. In this work we overcome these challenges through the coupling of overs ling MALDI-MSI with laser post-ionisation (MALDI-2). We demonstrate the ability to acquire rich lipid spectra from pixels as small as 6 μm, equivalent to or smaller than the size of typical mammalian cells. Coupled with an approach for automated lipid identification, it is shown that MALDI-2 combined with overs ling at 6 μm pixel size can detect up to three times more lipids and many more lipid classes than even conventional MALDI at 20 μm resolution in the positive-ion mode. Applying this to mouse kidney and human brain tissue containing active multiple sclerosis lesions, where 74 and 147 unique lipids are identified, respectively, the localisation of lipid signals to in idual tubuli within the kidney and lipid droplets with lesion-specific macrophages is demonstrated.
Publisher: American Chemical Society (ACS)
Date: 21-11-2019
Publisher: American Chemical Society (ACS)
Date: 16-01-2020
Publisher: Cold Spring Harbor Laboratory
Date: 28-08-2023
DOI: 10.1101/2023.08.28.555056
Abstract: Recent advances in spatial omics methods are revolutionising biomedical research by enabling detailed molecular analyses of cells and their interactions in their native state. As most technologies capture only a specific type of molecules, there is an unmet need to enable integration of multiple spatial-omics datasets. This, however, presents several challenges as these analyses typically operate on separate tissue sections at disparate spatial resolutions. Here, we established a spatial multi-omics integration pipeline enabling co-registration and granularity matching, and applied it to integrate spatial transcriptomics, mass spectrometry-based lipidomics, single nucleus RNA-seq and histomorphological information from human prostate cancer patient s les. This approach revealed unique correlations between lipids and gene expression profiles that are linked to distinct cell populations and histopathological disease states and uncovered molecularly different subregions not discernible by morphology alone. By its ability to correlate datasets that span across the biomolecular and spatial scale, the application of this novel spatial multi-omics integration pipeline provides unprecedented insight into the intricate interplay between different classes of molecules in a tissue context. In addition, it has unique hypothesis-generating potential, and holds promise for applications in molecular pathology, biomarker and target discovery and other tissue-based research fields.
Publisher: Cold Spring Harbor Laboratory
Date: 07-10-2020
DOI: 10.1101/2020.10.07.330530
Abstract: Mass spectrometry imaging (MSI) visualises molecular distributions throughout tissues but is blind to dynamic metabolic processes. Here, MSI with high mass resolution together with multiple stable isotope labelling provided spatial analyses of phosphatidylcholine (PC) metabolism in mouse lungs. Dysregulated surfactant metabolism is central to many respiratory diseases. Metabolism and turnover of therapeutic pulmonary surfactants were imaged from distributions of intact and metabolic products of an added tracer, universally 13 C-labelled dipalmitoyl PC (U[ 13 C]DPPC). The parenchymal distributions of newly synthesised PC species were also imaged from incorporations of methyl -D 9 -choline. This dual labelling strategy demonstrated both lack of inhibition of endogenous PC synthesis by exogenous surfactant and location of acyl chain remodelling processes acting on the U[ 13 C]DPPC-labelled surfactant, leading to formation of polyunsaturated PC lipids. This ability to visualise discrete metabolic events will greatly enhance our understanding of lipid metabolism in erse tissues, and has potential application to both clinical and experimental studies.
Publisher: Oxford University Press (OUP)
Date: 08-2015
Publisher: American Chemical Society (ACS)
Date: 24-05-2016
DOI: 10.1021/ACS.ANALCHEM.6B00672
Abstract: Mass spectrometry imaging (MSI) is a powerful molecular imaging technique. In microprobe MSI, images are created through a grid-wise interrogation of in idual spots by mass spectrometry across a surface. Classical statistical tests for within-s le comparisons fail as close-by measurement spots violate the assumption of independence of these tests, which can lead to an increased false-discovery rate. For spatial data, this effect is referred to as spatial autocorrelation. In this study, we investigated spatial autocorrelation in three different matrix-assisted laser desorption/ionization MSI data sets. These data sets cover different molecular classes (metabolites/drugs, lipids, and proteins) and different spatial resolutions ranging from 20 to 100 μm. Significant spatial autocorrelation was detected in all three data sets and found to increase with decreasing pixel size. To enable statistical testing for differences in mass signal intensities between regions of interest within MSI data sets, we propose the use of Conditional Autoregressive (CAR) models. We show that, by accounting for spatial autocorrelation, discovery rates (i.e., the ratio between the features identified and the total number of features) could be reduced between 21% and 69%. The reliability of this approach was validated by control mass signals based on prior knowledge. In light of the advent of larger MSI data sets based on either an increased spatial resolution or 3D data sets, accounting for effects due to spatial autocorrelation becomes even more indispensable. Here, we propose a generic and easily applicable workflow to enable within-s le statistical comparisons.
Publisher: American Chemical Society (ACS)
Date: 06-12-2022
Publisher: Elsevier BV
Date: 02-2018
DOI: 10.1194/JLR.M080093
Publisher: Elsevier BV
Date: 10-2013
DOI: 10.1016/J.PLIPRES.2013.04.005
Abstract: Mass spectrometry is now an indispensable tool for lipid analysis and is arguably the driving force in the renaissance of lipid research. In its various forms, mass spectrometry is uniquely capable of resolving the extensive compositional and structural ersity of lipids in biological systems. Furthermore, it provides the ability to accurately quantify molecular-level changes in lipid populations associated with changes in metabolism and environment bringing lipid science to the "omics" age. The recent explosion of mass spectrometry-based surface analysis techniques is fuelling further expansion of the lipidomics field. This is evidenced by the numerous papers published on the subject of mass spectrometric imaging of lipids in recent years. While imaging mass spectrometry provides new and exciting possibilities, it is but one of the many opportunities direct surface analysis offers the lipid researcher. In this review we describe the current state-of-the-art in the direct surface analysis of lipids with a focus on tissue sections, intact cells and thin-layer chromatography substrates. The suitability of these different approaches towards analysis of the major lipid classes along with their current and potential applications in the field of lipid analysis are evaluated.
Publisher: Elsevier BV
Date: 2021
Publisher: American Chemical Society (ACS)
Date: 29-07-2019
Publisher: American Chemical Society (ACS)
Date: 15-06-2023
Publisher: Elsevier BV
Date: 08-2018
DOI: 10.1016/J.BBADIS.2018.04.022
Abstract: Acute viral myocarditis (VM), characterised by leukocyte infiltration and dysfunction of the heart, is an important cause of sudden cardiac death in young adults. Unfortunately, to date, the pathological mechanisms underlying cardiac failure in VM remain incompletely understood. In the current study, we investigated if acute VM leads to cardiac metabolic rewiring and if this process is driven by local inflammation. Transcriptomic analysis of cardiac biopsies from myocarditis patients and a mouse model of VM revealed prominent reductions in the expression of a multitude of genes involved in mitochondrial oxidative energy metabolism. In mice, this coincided with reductions in high-energy phosphate and NAD levels, as determined by Imaging Mass Spectrometry, as well as marked decreases in the activity, protein abundance and mRNA levels of various enzymes and key regulators of cardiac oxidative metabolism. Indicative of fulminant cardiac inflammation, NF-κB signalling and inflammatory cytokine expression were potently induced in the heart during human and mouse VM. In cultured cardiomyocytes, cytokine-mediated NF-κB activation impaired cardiomyocyte oxidative gene expression, likely by interfering with the PGC-1 (peroxisome proliferator-activated receptor (PPAR)-γ co-activator) signalling network, the key regulatory pathway controlling cardiomyocyte oxidative metabolism. In conclusion, we provide evidence that acute VM is associated with extensive cardiac metabolic remodelling and our data support a mechanism whereby cytokines secreted primarily from infiltrating leukocytes activate NF-κB signalling in cardiomyocytes thereby inhibiting the transcriptional activity of the PGC-1 network and consequently modulating myocardial energy metabolism.
Publisher: Springer Science and Business Media LLC
Date: 08-08-2022
Publisher: Elsevier BV
Date: 04-2014
Publisher: Springer Science and Business Media LLC
Date: 18-02-2019
DOI: 10.1038/S41598-018-38257-0
Abstract: Treatment for medulloblastoma (MB) — the most common malignant pediatric brain tumor — includes prophylactic radiation administered to the entire brain and spine due to the high incidence of metastasis to the central nervous system. However, the majority of long-term survivors are left with permanent and debilitating neurocognitive impairments as a result of this therapy, while the remaining 30–40% of patients relapse with terminal metastatic disease. Development of more effective targeted therapies has been hindered by our lack of understanding of the underlying mechanisms regulating the metastatic process in this disease. To understand the mechanism by which MB metastasis occurs, three-dimensional matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) experiments were performed on whole brains from a mouse model of human medulloblastoma. Analyzing the tumor and surrounding normal brain in its entirety enabled the detection of low abundance, spatially-heterogeneous lipids associated with tumor development. Boundaries of metastasizing and non-metastasizing primary tumors were readily defined, leading to the identification of lipids associated with medulloblastoma metastasis, including phosphatidic acids, phosphatidylethanolamines, phosphatidylserines, and phosphoinositides. These lipids provide a greater insight into the metastatic process and may ultimately lead to the discovery of biomarkers and novel targets for the diagnosis and treatment of metastasizing MB in humans.
Publisher: American Association for the Advancement of Science (AAAS)
Date: 15-04-2022
Abstract: Human cells produce thousands of lipids that change during cell differentiation and can vary across in idual cells of the same type. However, we are only starting to characterize the function of these cell-to-cell differences in lipid composition. Here, we measured the lipidomes and transcriptomes of in idual human dermal fibroblasts by coupling high-resolution mass spectrometry imaging with single-cell transcriptomics. We found that the cell-to-cell variations of specific lipid metabolic pathways contribute to the establishment of cell states involved in the organization of skin architecture. Sphingolipid composition is shown to define fibroblast subpopulations, with sphingolipid metabolic rewiring driving cell-state transitions. Therefore, cell-to-cell lipid heterogeneity affects the determination of cell states, adding a new regulatory component to the self-organization of multicellular systems.
Publisher: Wiley
Date: 26-10-2018
Abstract: Grain boundaries play a key role in the performance of thin-film optoelectronic devices and yet their effect in halide perovskite materials is still not understood. The biggest factor limiting progress is the inability to identify grain boundaries. Noncrystallographic techniques can misidentify grain boundaries, leading to conflicting literature reports about their influence however, the gold standard - electron backscatter diffraction (EBSD) - destroys halide perovskite thin films. Here, this problem is solved by using a solid-state EBSD detector with 6000 times higher sensitivity than the traditional phosphor screen and camera. Correlating true grain size with photoluminescence lifetime, carrier diffusion length, and mobility shows that grain boundaries are not benign but have a recombination velocity of 1670 cm s
Publisher: American Chemical Society (ACS)
Date: 07-11-2012
DOI: 10.1021/AC302634U
Abstract: The on-demand printing of living cells using inkjet technologies has recently been demonstrated and allows for the controlled deposition of cells in microarrays. Here, we show that such arrays can be interrogated directly by robot-controlled liquid microextraction coupled with chip-based nanoelectospray mass spectrometry. Such automated analyses generate a profile of abundant membrane lipids that are characteristic of cell type. Significantly, the spatial control in both deposition and extraction steps combined with the sensitivity of the mass spectrometric detection allows for robust molecular profiling of in idual cells.
Publisher: American Chemical Society (ACS)
Date: 28-06-2017
DOI: 10.1021/ACS.ANALCHEM.7B01168
Abstract: Matrix-Assisted Laser Desorption Ionization, MALDI, has been increasingly used in a variety of biomedical applications, including tissue imaging of clinical tissue s les, and in drug discovery and development. These studies strongly depend on the performance of the analytical instrumentation and would drastically benefit from improved sensitivity, reproducibility, and mass/spatial resolution. In this work, we report on a novel combined MALDI/ESI interface, which was coupled to different Orbitrap mass spectrometers (Elite and Q Exactive Plus) and extensively characterized with peptide and protein standards, and in tissue imaging experiments. In our approach, MALDI is performed in the elevated pressure regime (5-8 Torr) at a spatial resolution of 15-30 μm, while ESI-generated ions are injected orthogonally to the interface axis. We have found that introduction of the MALDI-generated ions into an electrodynamic dual-funnel interface results in increased sensitivity characterized by a limit of detection of ∼400 zmol, while providing a mass measurement accuracy of 1 ppm and a mass resolving power of 120 000 in analysis of protein digests. In tissue imaging experiments, the MALDI/ESI interface has been employed in experiments with rat brain sections and was shown to be capable of visualizing and spatially characterizing very low abundance analytes separated only by 20 mDa. Comparison of imaging data has revealed excellent agreement between the MALDI and histological images.
Publisher: Wiley
Date: 04-12-2021
Abstract: Matrix‐assisted laser/desorption ionisation‐mass spectrometry imaging (MALDI‐MSI) enables label‐free imaging of biomolecules in biological tissues. However, many molecules remain undetected due to their poor ionisation efficiencies. These poor ionisation efficiencies practically limit spatial resolution. Herein, we address this challenge for aromatic antioxidants by reporting an innovative approach involving sequential matrix‐assisted laser desorption and two‐photon ionisation of desorbed neutrals. It is shown that ion yields increase with reduced s ling areas obtained using sub‐threshold primarily laser fluence. This counterintuitive observation could arise from a reduction in radical/ion neutralisation reactions within the sparse plume and/or favorable molecular desorption under low fluence conditions. The utility of this approach is demonstrated for imaging tocopherols and ubiquinols in mouse brain and prostate cancer tissue. This can pave the way for improved sensitivity in MSI experiments at cellular and sub‐cellular resolutions.
Publisher: Elsevier BV
Date: 12-2019
Publisher: Wiley
Date: 11-05-2017
Publisher: Elsevier BV
Date: 09-2014
DOI: 10.1016/J.BBALIP.2014.05.006
Abstract: The lipidome of the human lens is unique in that cholesterol and dihydrosphingomyelin are the dominant classes. Moreover, the lens lipidome is not static with dramatic changes in several sphingolipid classes associated with both aging and cataract. Accordingly, there is a clear need to expand knowledge of the molecular species that constitute the human lens sphingolipidome. In this study, human lens lipids have been extracted and separated by thin-layer chromatography (TLC). Direct analysis of the TLC plates by desorption electrospray ionisation-mass spectrometry (DESI-MS) allowed the detection over 30 species from 11 classes of sphingolipids. Significantly, novel classes of lens lipids including sulfatides, dihydrosulfatides, lactosylceramide sulfates and dihydrolactosylceramide sulfates were identified.
Publisher: American Chemical Society (ACS)
Date: 11-01-2022
Abstract: Understanding how neutral molecules become protonated during positive-ion electrospray ionization (ESI) mass spectrometry is critically important to ensure analytes can be efficiently ionized, detected, and unambiguously identified. The ESI solvent is one of several parameters that can alter the dominant site of protonation in polyfunctional molecules and thus, in turn, can significantly change the collision-induced dissociation (CID) mass spectra relied upon for compound identification. Ciprofloxacin─a common fluoroquinolone antibiotic─is one such ex le whereby positive-ion ESI can result in gas-phase [M + H]
Publisher: Wiley
Date: 19-06-2018
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C7CC02325A
Abstract: Coupling laser post-ionisation with a high resolving power MALDI Orbitrap mass spectrometer has realised an up to ∼100-fold increase in the sensitivity and enhanced the chemical coverage for MALDI-MS imaging of lipids relative to conventional MALDI.
Publisher: American Chemical Society (ACS)
Date: 10-12-2013
DOI: 10.1021/AC403421V
Abstract: This letter demonstrates the use of infrared matrix-assisted laser desorption/ionization coupled with microscope mode mass spectrometry imaging. It is aimed to explore the use of intrinsic water in tissue as a matrix for imaging at spatial resolutions below the diffraction limit of the employed IR optics. Stigmatic ion optics with a magnification factor of ~70 were used to project the spatial distribution of produced ions onto a detector while separating ions with different mass-to-charge ratios using a time-of-flight mass spectrometer. A pixelated detector was used to simultaneously record arrival time and impact position. A previously described dried-droplet s le system of 2,5-dihydroxybenzoic acid (DHB) and 5 peptides covered by a copper grid for defined surface structure was used to benchmark the light- and ion-optical setup for spatial resolution and mass spectrometric performance. A spatial resolving power of 9.8 μm, well below the optical limit of diffraction (14 μm for the given setup), was established. After, frozen cryo-sections from a biological model system were measured by exploiting the endogenous water content as a matrix. Principal component analysis enabled a clear distinction between distinct tissue regions identified by both light microscopy and MS imaging.
Publisher: American Chemical Society (ACS)
Date: 08-05-2017
Publisher: Springer Science and Business Media LLC
Date: 27-12-2019
DOI: 10.1038/S41598-019-56364-4
Abstract: Mass spectrometry imaging (MSI) is a technique for mapping the spatial distributions of molecules in sectioned tissue. Histology-preserving tissue preparation methods are central to successful MSI studies. Common fixation methods, used to preserve tissue morphology, can result in artifacts in the resulting MSI experiment including delocalization of analytes, altered adduct profiles, and loss of key analytes due to irreversible cross-linking and diffusion. This is especially troublesome in lung and airway s les, in which histology and morphology is best interpreted from 3D reconstruction, requiring the large and small airways to remain inflated during analysis. Here, we developed an MSI-compatible inflation containing as few exogenous components as possible, forgoing perfusion, fixation, and addition of salt solutions upon inflation that resulted in an ungapped 3D molecular reconstruction through more than 300 microns. We characterized a series of polyunsaturated phospholipids (PUFA-PLs), specifically phosphatidylinositol (-PI) lipids linked to lethal inflammation in bacterial infection and mapped them in serial sections of inflated mouse lung. PUFA-PIs were identified using spatial lipidomics and determined to be determinant markers of major airway features using unsupervised hierarchical clustering. Deep lung architecture was preserved using this inflation approach and the resulting sections are compatible with multiple MSI modalities, automated interpretation software, and serial 3D reconstruction.
Publisher: Elsevier BV
Date: 11-2018
Publisher: Cold Spring Harbor Laboratory
Date: 22-06-2023
DOI: 10.1101/2023.06.21.546027
Abstract: Mass spectrometry imaging (MSI) has accelerated the understanding of lipid metabolism and spatial distribution in tissues and cells. However, few MSI studies have approached lipid imaging quantitatively and those that have focus on a single lipid class. Herein, we overcome limitation of quantitative MSI (Q-MSI) by using a multi-class internal standard lipid mixture that is sprayed homogenously over the tissue surface with analytical concentrations that reflects endogenous brain lipid levels. Using this approach we have performed Q-MSI for 13 lipid classes representing sum-composition lipid species. This was carried out using both MALDI (negative ion mode) and MALDI-2 (positive ion mode) and pixel-wise normalisation of each lipid species signal to the corresponding class-specific IS an approach analogous to that widely used for shotgun lipidomics from biological extracts. This approach allows pixel concentrations of lipids to be reported in pmol/mm 2 . Q-MSI of lipids covered 3 orders of magnitude in dynamic range and revealed subtle change sin in distribution compared to conventional total-ion-current normalisation approaches. The robustness of the method was evaluated by repeating experiments in two laboratories on biological replicates using both timsTOF and Orbitrap mass spectrometers operated with a ~4-fold difference in mass resolution power. There was a strong overall correlation in the Q-MSI result obtained using the two approaches with outliers mostly rationalised by isobaric interferences that are only resolved with the Orbitrap system or the higher sensitivity of one instrument for particular lipid species, particularly for lipids detected at low intensity. These data provide insight into how mass resolving power can affect Q-MSI data. This approach opens up the possibility of performing large-scale Q-MSI studies across numerous lipid classes and reveal how absolute lipid concentrations vary throughout and between biological tissues.
Publisher: Wiley
Date: 18-02-2019
Publisher: Springer Science and Business Media LLC
Date: 09-01-2019
DOI: 10.1038/S41598-018-36957-1
Abstract: Desorption electrospray ionisation-mass spectrometry imaging (DESI-MSI) is a powerful imaging technique for the analysis of complex surfaces. However, the often highly complex nature of biological s les is particularly challenging for MSI approaches, as options to appropriately address molecular complexity are limited. Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) offers superior mass accuracy and mass resolving power, but its moderate throughput inhibits broader application. Here we demonstrate the dramatic gains in mass resolution and/or throughput of DESI-MSI on an FT-ICR MS by developing and implementing a sophisticated data acquisition and data processing pipeline. The presented pipeline integrates, for the first time, parallel ion accumulation and detection, post-processing absorption mode Fourier transform and pixel-by-pixel internal re-calibration. To achieve that, first, we developed and coupled an external high-performance data acquisition system to an FT-ICR MS instrument to record the time-domain signals (transients) in parallel with the instrument’s built-in electronics. The recorded transients were then processed by the in-house developed computationally-efficient data processing and data analysis software. Importantly, the described pipeline is shown to be applicable even to extremely large, up to 1 TB, imaging datasets. Overall, this approach provides improved analytical figures of merits such as: (i) enhanced mass resolution at no cost in experimental time and (ii) up to 4-fold higher throughput while maintaining a constant mass resolution. Using this approach, we not only demonstrate the record 1 million mass resolution for lipid imaging from brain tissue, but explicitly show such mass resolution is required to resolve the complexity of the lipidome.
Publisher: American Chemical Society (ACS)
Date: 12-2010
DOI: 10.1016/J.JASMS.2010.09.003
Abstract: The lipid composition of the human lens is distinct from most other tissues in that it is high in dihydrosphingomyelin and the most abundant glycerophospholipids in the lens are unusual 1-O-alkyl-ether linked phosphatidylethanolamines and phosphatidylserines. In this study, desorption electrospray ionization (DESI) mass spectrometry-imaging was used to determine the distribution of these lipids in the human lens along with other lipids including, ceramides, ceramide-1-phosphates, and lyso 1-O-alkyl ethers. To achieve this, 25 μm lens slices were mounted onto glass slides and analyzed using a linear ion-trap mass spectrometer equipped with a custom-built, 2-D automated DESI source. In contrast to other tissues that have been previously analyzed by DESI, the presence of a strong acid in the spray solvent was required to desorb lipids directly from lens tissue. Distinctive distributions were observed for [M + H](+) ions arising from each lipid class. Of particular interest were ionized 1-O-alkyl phosphatidylethanolamines and phosphatidylserines, PE (18:1e/18:1), and PS (18:1e/18:1), which were found in a thin ring in the outermost region of the lens. This distribution was confirmed by quantitative analysis of lenses that were sectioned into four distinct regions (outer, barrier, inner, and core), extracted and analyzed by electrospray ionization tandem mass spectrometry. DESI-imaging also revealed a complementary distribution for the structurally-related lyso 1-O-alkyl phosphatidylethanolamine, LPE (18:1e), which was localized closer to the centre of the lens. The data obtained in this study indicate that DESI-imaging is a powerful tool for determining the spatial distribution of human lens lipids.
Publisher: American Chemical Society (ACS)
Date: 19-01-2020
Publisher: American Chemical Society (ACS)
Date: 15-08-2023
Publisher: Wiley
Date: 19-06-2018
Publisher: Springer Science and Business Media LLC
Date: 27-11-2013
DOI: 10.1007/S00216-013-7478-9
Abstract: Mass spectrometry imaging (MSI) has evolved into a valuable tool across many fields of chemistry, biology, and medicine. However, arguably its greatest disadvantage is the difficulty in acquiring quantitative data regarding the surface concentration of the analyte(s) of interest. These difficulties largely arise from the high dependence of the ion signal on the localized chemical and morphological environment and the difficulties associated with calibrating such signals. The development of quantitative MSI approaches would correspond to a giant leap forward for the field, particularly for the biomedical and pharmaceutical fields, and is thus a highly active area of current research. In this review, we outline the current progress being made in the development and application of quantitative MSI workflows with a focus on biomedical applications. Particular emphasis is placed on the various strategies used for both signal calibration and correcting for various ion suppression effects that are invariably present in any MSI study. In addition, the difficulties in validating quantitative-MSI data on a pixel-by-pixel basis are highlighted.
Publisher: Frontiers Media SA
Date: 05-08-2021
DOI: 10.3389/FENDO.2021.689600
Abstract: Prostate cancer is the fourth most common cancer worldwide with definitive diagnosis reliant on biopsy and human-graded histopathology. As with other pathologies, grading based on classical haematoxylin and eosin (H& E) staining of formalin fixed paraffin-embedded material can be prone to variation between pathologists, prompting investigation of biomolecular markers. Comprising around 50% of cellular mass, and with known metabolic variations in cancer, lipids provide a promising target for molecular pathology. Here we apply isomer-resolved lipidomics in combination with imaging mass spectrometry to interrogate tissue sections from radical prostatectomy specimens. Guided by the histopathological assessment of adjacent tissue sections, regions of interest are investigated for molecular signatures associated with lipid metabolism, especially desaturation and elongation pathways. Monitoring one of the most abundant cellular membrane lipids within these tissues, phosphatidylcholine (PC) 34:1, high positive correlation was observed between the n-9 isomer (site of unsaturation 9-carbons from the methyl terminus) and epithelial cells from potential pre-malignant lesions, while the n-7 isomer abundance was observed to correlate with immune cell infiltration and inflammation. The correlation of lipid isomer signatures with human disease states in tissue suggests a future role for isomer-resolved mass spectrometry imaging in assisting pathologists with prostate cancer diagnoses and patient stratification.
Publisher: American Chemical Society (ACS)
Date: 22-03-2014
DOI: 10.1007/S13361-014-0839-6
Abstract: In this study, we describe the implementation of a position- and time-sensitive detection system (Timepix detector) to directly visualize the spatial distributions of the matrix-assisted laser desorption ionization ion cloud in a linear-time-of-flight (MALDI linear-ToF) as it is projected onto the detector surface. These time-resolved images allow direct visualization of m/z-dependent ion focusing effects that occur within the ion source of the instrument. The influence of key parameters, namely extraction voltage (E(V)), pulsed-ion extraction (PIE) delay, and even the matrix-dependent initial ion velocity was investigated and were found to alter the focusing properties of the ion-optical system. Under certain conditions where the spatial focal plane coincides with the detector plane, so-called x-y space focusing could be observed (i.e., the focusing of the ion cloud to a small, well-defined spot on the detector). Such conditions allow for the stigmatic ion imaging of intact proteins for the first time on a commercial linear ToF-MS system. In combination with the ion-optical magnification of the system (~100×), a spatial resolving power of 11–16 μm with a pixel size of 550 nm was recorded within a laser spot diameter of ~125 μm. This study demonstrates both the diagnostic and analytical advantages offered by the Timepix detector in ToF-MS.
Publisher: eLife Sciences Publications, Ltd
Date: 11-03-2014
DOI: 10.7554/ELIFE.01751
Abstract: Pheromones play an important role in the behavior, ecology, and evolution of many organisms. The structure of many insect pheromones typically consists of a hydrocarbon backbone, occasionally modified with various functional oxygen groups. Here we show that sex-specific triacylclyerides (TAGs) are broadly conserved across the subgenus Drosophila in 11 species and represent a novel class of pheromones that has been largely overlooked. In desert-adapted drosophilids, 13 different TAGs are secreted exclusively by males from the ejaculatory bulb, transferred to females during mating, and function synergistically to inhibit courtship from other males. Sex-specific TAGs are comprised of at least one short branched tiglic acid and a long linear fatty acyl component, an unusual structural motif that has not been reported before in other natural products. The ersification of chemical cues used by desert-adapted Drosophila as pheromones may be related to their specialized diet of fermenting cacti.
Publisher: Royal Society of Chemistry (RSC)
Date: 2016
DOI: 10.1039/C6AN00169F
Abstract: Here, we reveal the increased biochemical and spatial information acquired using high-speed MALDI-MSI and sequential acquisitions of positive and negative lipid-MSI data from single tissue sections.
Publisher: Springer Science and Business Media LLC
Date: 06-09-2011
Publisher: Royal Society of Chemistry (RSC)
Date: 2012
DOI: 10.1039/C1AN15864C
Abstract: Unsaturated lipids deposited onto a range of materials are observed to react with the low concentrations of ozone present in normal laboratory air. Parent lipids and ozonolysis cleavage products are both detected directly from surfaces by desorption electrospray ionisation mass spectrometry (DESI-MS) with the resulting mass spectra providing clear evidence of the double bond position within these molecules. This serendipitous process has been coupled with thin-layer chromatography (TLC) to provide a simple but powerful approach for the detailed structural elucidation of lipids present in complex biological extracts. Lipid extracts from human lens were deposited onto normal phase TLC plates and then developed to separate components according to lipid class. Exposure of the developed plates to laboratory air for ca. 1 h prior to DESI-MS analysis gave rise to ozonolysis products allowing for the unambiguous identification of double bond positions in even low abundant, unsaturated lipids. In particular, the co-localization of intact unsaturated lactosylceramides (LacCer) with products from their oxidative cleavage provide the first evidence for the presence of three isomeric LacCer (d18:0/24:1) species in the ocular lens lipidome, i.e., variants with double bonds at the n-9, n-7 and n-5 positions.
Publisher: American Chemical Society (ACS)
Date: 06-07-2021
Publisher: Rockefeller University Press
Date: 25-02-2020
DOI: 10.1084/JEM.20191660
Abstract: Failure of remyelination underlies the progressive nature of demyelinating diseases such as multiple sclerosis. Macrophages and microglia are crucially involved in the formation and repair of demyelinated lesions. Here we show that myelin uptake temporarily skewed these phagocytes toward a disease-resolving phenotype, while sustained intracellular accumulation of myelin induced a lesion-promoting phenotype. This phenotypic shift was controlled by stearoyl-CoA desaturase-1 (SCD1), an enzyme responsible for the desaturation of saturated fatty acids. Monounsaturated fatty acids generated by SCD1 reduced the surface abundance of the cholesterol efflux transporter ABCA1, which in turn promoted lipid accumulation and induced an inflammatory phagocyte phenotype. Pharmacological inhibition or phagocyte-specific deficiency of Scd1 accelerated remyelination ex vivo and in vivo. These findings identify SCD1 as a novel therapeutic target to promote remyelination.
Publisher: American Chemical Society (ACS)
Date: 23-03-2015
DOI: 10.1021/AC5041764
Abstract: A key requirement of electrospray ionization (ESI) and other techniques facilitating ionization at elevated pressures is the efficient transport of free gas-phase ions into the high vacuum region of the mass spectrometer. Radio frequency (RF) multipole ion guides that allow for collisional cooling are one of the most popular means of achieving this. However, their performance is highly dependent on several experimental factors, including pressure and various electrode potentials along the ion path. To experimentally visualize these effects, we have employed a position-sensitive detector at the exit of a quadrupole mass spectrometer (QMS) instrument operated in RF only mode that employs an RF only octopole as a collisional cooling ion guide. This allows the spatial distribution of the ions, and its dependence on experimentally determined conditions, to be directly visualized at the exit of the quadrupole. This investigation provides a detailed insight into the ion dynamics occurring inside multipole ion guides. This knowledge can directly be applied to instrument development and to improve the ion transmission efficiency and, thus, sensitivity. Numerical simulations using custom-developed trajectory simulation software are compared and contrasted with the experimental observations.
Publisher: Springer Science and Business Media LLC
Date: 21-05-2018
DOI: 10.1038/S41592-018-0010-6
Abstract: We report a method that enables automated data-dependent acquisition of lipid tandem mass spectrometry data in parallel with a high-resolution mass spectrometry imaging experiment. The method does not increase the total image acquisition time and is combined with automatic structural assignments. This lipidome-per-pixel approach automatically identified and validated 104 unique molecular lipids and their spatial locations from rat cerebellar tissue.
Publisher: Wiley
Date: 18-02-2019
Publisher: American Chemical Society (ACS)
Date: 22-10-2018
Publisher: Royal Society of Chemistry (RSC)
Date: 2017
DOI: 10.1039/C7TB01100H
Abstract: A review highlighting current trends, key applications, and the vast potential of mass spectrometry imaging for biomaterials research.
Publisher: Wiley
Date: 26-01-2021
Publisher: American Chemical Society (ACS)
Date: 12-08-2014
DOI: 10.1007/S13361-014-0958-0
Abstract: In this paper, we have employed an ion imaging approach to investigate the behavior of ions exiting from a quadrupole mass spectrometer (QMS) system that employs a radio frequency octopole ion guide before the QMS. An in-vacuum active pixel detector (Timepix) is employed at the exit of the QMS to image the ion patterns. The detector assembly simultaneously records the ion impact position and number of ions per pixel in every measurement frame. The transmission characteristics of the ion beam exiting the QMS are studied using this imaging detector under different operating conditions. Experimental results confirm that the ion spatial distribution exiting the QMS is heavily influenced by ion injection conditions. Furthermore, ion images from Timepix measurements of protein standards demonstrate the capability to enhance the quality of the mass spectral information and provide a detailed insight in the spatial distribution of different charge states (and hence different m/z) ions exiting the QMS.
Publisher: American Chemical Society (ACS)
Date: 05-07-2018
Publisher: Elsevier BV
Date: 08-2019
Publisher: Wiley
Date: 17-02-2021
Publisher: Wiley
Date: 28-03-2022
DOI: 10.1002/JMS.4820
Abstract: Time‐of‐flight (TOF) systems are one of the most widely used mass analyzers in native mass spectrometry (nMS) for the analysis of non‐covalent multiply charged bio‐macromolecular assemblies (MMAs). Typically, microchannel plates (MCPs) are employed for high mass native ion detection in TOF MS. MCPs are well known for their reduced detection efficiency when impinged by large slow moving ions. Here, a position‐ and time‐sensitive Timepix (TPX) detector has been added to the back of a dual MCP stack to study the key factors that affect MCP performance for MMA ions generated by nMS. The footprint size of the secondary electron cloud generated by the MCP on the TPX for each in idual ion event is analyzed as a measure of MCP performance at each mass‐to‐charge ( m/z ) value and resulted in a Poisson distribution. This allowed us to investigate the dependency of ion mass, ion charge, ion velocity, acceleration voltage, and MCP bias voltage on MCP response in the high mass low velocity regime. The study of measurement ranges ion mass = 195 to 802,000 Da, ion velocity = 8.4 to 67.4 km/s, and ion charge = 1+ to 72+, extended the previously examined mass range and characterized MCP performance for multiply charged species. We derived a MCP performance equation based on two independent ion properties, ion mass and charge, from these results, which enables rapid MCP tuning for single MMA ion detection.
Publisher: Wiley
Date: 28-10-2015
DOI: 10.1002/RCM.7379
Abstract: In mass spectrometry imaging (MSI) it is often desirable to analyse the same s le in both polarities to extract the most information. However, many matrices that produce high-quality spectra in matrix-assisted laser desorption/ionization (MALDI) are volatile, greatly limiting their use in long imaging experiments. We demonstrate that using a new high speed MALDI-MSI instrument, volatile matrices, including those that produce intense lipid signals in both positive and negative ion mode, can now be effectively used in MSI. A prototype Bruker rapifleX MALDI Tissuetyper™ time-of-flight (TOF) instrument was used for high-speed imaging. This allows acquisition rates up to 50 pixels/s made possible by use of a 10 kHz laser and two rotating mirrors that allow the laser beam to be moved over, and synchronised with, the rapidly moving s le. MSI experiments were performed on mouse brain sections using non-vacuum stable dithranol and 2,6-dihydroxyacetophenone (DHA) matrices with pixel sizes ranging from 10 × 10 µm(2) to 50 × 50 µm(2). Both DHA and dithranol produced rich, complementary lipid spectra in both positive and negative ion modes. Due to the rapid acquisition speed of the instrument, both matrices could be effectively used for MSI despite their volatility. For ex le, an entire mouse brain could be imaged consecutively in both positive and negative ion mode with 50 × 50 µm(2) pixels in ~35 min. We demonstrate that these speeds make possible both faster and higher resolution imaging of biological tissues on practical timescales. These high acquisition speeds now make possible whole new classes of matrices that are unstable under high vacuum for MALDI-MSI studies. This provides researchers with far greater range and flexibility in choosing the best matrix for the given s le and analytes that they wish to detect. In addition, such instruments allow MSI to be performed at higher resolution across larger areas on practical time scales.
Publisher: American Chemical Society (ACS)
Date: 09-03-2023
Publisher: Wiley
Date: 03-09-2013
Abstract: Flying high: Application of an active pixel detector with high charge sensitivity to a linear time-of-flight mass spectrometer results in enhanced detection of high-mass proteins (such as Immunoglobulin G IgG) using a conventional microchannel plate detection system. This technique thus provides a means to extend the mass range of such detectors as well as allowing direct visualization of mass-dependent ion-focusing phenomena.
Publisher: Elsevier BV
Date: 04-2019
Publisher: Royal Society of Chemistry (RSC)
Date: 2023
DOI: 10.1039/D3AY01046E
Abstract: Laser post-ionisation coupled with MALDI-mass spectrometry imaging significantly enhances the sensitivity and coverage of small metabolites.
Publisher: Elsevier BV
Date: 02-2021
DOI: 10.1016/J.CELREP.2021.108738
Abstract: Canonical fatty acid metabolism describes specific enzyme-substrate interactions that result in products with well-defined chain lengths, degree(s), and positions of unsaturation. Deep profiling of lipids across a range of prostate cancer cell lines reveals a variety of fatty acids with unusual site(s) of unsaturation that are not described by canonical pathways. The structure and abundance of these unusual lipids correlate with changes in desaturase expression and are strong indicators of cellular phenotype. Gene silencing and stable isotope tracing demonstrate that direct Δ6 and Δ8 desaturation of 14:0 (myristic), 16:0 (palmitic), and 18:0 (stearic) acids by FADS2 generate new families of unsaturated fatty acids (including n-8, n-10, and n-12) that have rarely-if ever-been reported in human-derived cells. Isomer-resolved lipidomics reveals the selective incorporation of these unusual fatty acids into complex structural lipids and identifies their presence in cancer tissues, indicating functional roles in membrane structure and signaling.
Publisher: Elsevier BV
Date: 07-2021
Publisher: American Chemical Society (ACS)
Date: 27-12-2022
Publisher: Springer Science and Business Media LLC
Date: 06-10-2020
DOI: 10.1007/S11307-020-01541-5
Abstract: Mass spectrometry imaging (MSI) enables the visualization of molecular distributions on complex surfaces. It has been extensively used in the field of biomedical research to investigate healthy and diseased tissues. Most of the MSI studies are conducted in a 2D fashion where only a single slice of the full s le volume is investigated. However, biological processes occur within a tissue volume and would ideally be investigated as a whole to gain a more comprehensive understanding of the spatial and molecular complexity of biological s les such as tissues and cells. Mass spectrometry imaging has therefore been expanded to the 3D realm whereby molecular distributions within a 3D s le can be visualized. The benefit of investigating volumetric data has led to a quick rise in the application of single-s le 3D-MSI investigations. Several experimental and data analysis aspects need to be considered to perform successful 3D-MSI studies. In this review, we discuss these aspects as well as ongoing developments that enable 3D-MSI to be routinely applied to multi-s le studies.
Publisher: American Chemical Society (ACS)
Date: 23-03-2018
Publisher: Cold Spring Harbor Laboratory
Date: 03-11-2021
DOI: 10.1101/2021.11.01.466716
Abstract: 1.0 Cellular energy and biomass demands of cancer drive a complex dynamic between uptake of extracellular fatty acids (FA) and de novo synthesis. Given that oxidation of de novo synthesised FAs for energy would result in net-energy loss, there is an implication that FAs from these two sources must have distinct metabolic fates - however hitherto FAs were considered part of a common pool. To probe FA metabolic partitioning, cancer cells were supplemented with stable-isotope labelled FAs. Structural analysis of the resulting glycerophospholipids revealed that labelled FAs from uptake were largely incorporated to canonical ( sn -)positions on the glycerol backbone. Surprisingly, labelled FA uptake disrupted canonical isomer patterns of the unlabelled lipidome and induced repartitioning of n -3 and n -6 polyunsaturated-FAs into glycerophospholipid classes. These structural changes evidence differences in the metabolic fate of FAs derived from uptake or de novo sources and demonstrate unique signalling and remodelling behaviours usually hidden to conventional lipidomics. Lipid isomers reveal discrete metabolic compartmentalisation in cancer FAs derived from uptake and de novo synthesis have different metabolic fates Stearate uptake signals for PUFA ( n -3 and n -6) repartitioning between lipid classes sn -positional isomers are a marker for aberrant lipid metabolism
Publisher: Elsevier BV
Date: 02-2019
DOI: 10.1016/J.JPROT.2018.10.008
Abstract: Mass spectrometry imaging (MSI) has emerged as a powerful tool in biomedical research to reveal the localization of a broad scale of compounds ranging from metabolites to proteins in diseased tissues, such as malignant tumors. MSI is most commonly used for the two-dimensional imaging of tissues from multiple patients or for the three-dimensional (3D) imaging of tissue from a single patient. These applications are potentially introducing a s ling bias on a s le or patient level, respectively. The aim of this study is therefore to investigate the consequences of s ling bias on s le representativeness and on the precision of biomarker discovery for histological grading of human bladder cancers by MSI. We therefore submitted formalin-fixed paraffin-embedded tissues from 14 bladder cancer patients with varying histological grades to 3D analysis by matrix-assisted laser desorption/ionization (MALDI) MSI. We found that, after removing 20% of the data based on novel outlier detection routines for 3D-MSI data based on the evaluation of digestion efficacy and z-directed regression, on average 33% of a s le has to be measured in order to obtain sufficient coverage of the existing biological variance within a tissue s le. SIGNIFICANCE: In this study, 3D MALDI-MSI is applied for the first time on a cohort of bladder cancer patients using formalin-fixed paraffin-embedded (FFPE) tissue of bladder cancer resections. This work portrays the reproducibility that can be achieved when employing an optimized s le preparation and subsequent data evaluation approach. Our data shows the influence of s ling bias on the variability of the results, especially for a small patient cohort. Furthermore, the presented data analysis workflow can be used by others as a 3D FFPE data-analysis pipeline working on multi-patient 3D-MSI studies.
Publisher: American Chemical Society (ACS)
Date: 31-01-2019
Publisher: IOP Publishing
Date: 23-04-2014
Start Date: 01-2020
End Date: 05-2024
Amount: $734,162.00
Funder: Australian Research Council
View Funded ActivityStart Date: 09-2019
End Date: 09-2022
Amount: $293,985.00
Funder: Australian Research Council
View Funded ActivityStart Date: 06-2022
End Date: 05-2023
Amount: $630,880.00
Funder: Australian Research Council
View Funded Activity