Molecular MALDI imaging: an emerging technology for neuroscience studies

Mass spectrometry (MS) has become an essential tool for the detection, identification, and characterization of the molecular components of biological processes, such as those responsible for the dynamic properties of the nervous system. Generally, the application of these powerful techniques requires the destruction of the specimen under study, but recent technological advances have made it possible to apply the matrix-assisted laser desorption/ionization (MALDI) MS technique directly to tissue sections. The major advantage of direct MALDI analysis is that it enables the acquisition of local molecular expression profiles, while maintaining the topographic integrity of the tissue and avoiding time-consuming extraction, purification, and separation steps, which have the potential for introducing artifacts. With automation and the ability to display complex spectral data using imaging software, it is now possible to create multiple 2D maps of selected biomolecules in register with tissue sections, a method now known as MALDI Imaging, or MSI (for Mass Spectrometry Imaging). This creates, for example, an opportunity to correlate functional states, determined a priori with live recording or imaging, with the corresponding molecular maps obtained at the time the tissue is frozen and analyzed with MSI. We review the increasing application of MALDI Imaging to the analysis of molecular distributions of proteins and peptides in nervous tissues of both vertebrates and invertebrates, focusing in particular on recent studies of neurodegenerative diseases and early efforts to implement assays of neuronal development.     

Quality will determine the future of mass spectrometry imaging in clinical laboratories: the need for standardization

Introduction: Mass Spectrometry Imaging (MSI) provides information about the spatial distributions of different analytes on tissue sections and requires no homogenization techniques or labeling. It has a wide spectrum of applications ranging from biomarker discovery to drug response studies. MSI can be adapted to the clinical settings due to its ability to combine mass spectrometry and histological data.

Areas covered: The major obstacle to adapt MSI into clinical settings arises from the limited standardization in MSI experiments. We aimed to review the multi-site studies in MSI specifically focusing on reproducibility. Additionally, we emphasized the necessity of quality assessments in MSI for its potential usage in clinical settings.

Expert opinion: We discuss how important it is to follow optimized and standardized workflows in MSI and conduct potential quality assessments at important stages in order to adapt MSI into clinical applications.     

Mass spectrometry imaging: An emerging technology for the analysis of metabolites in insects

Mass spectrometry imaging (MSI) can visualize the composition, abundance, and spatial distribution of molecules in tissues or cells, which has been widely used in the research of life science. Insects, especially the agricultural pests, have received a great deal of interests from the scientists in biodiversity and food security. This review introduces the major characteristics of MSI, summarizes its application to the investigation of insect endogenous metabolites, exogenous metabolites, and the spatiotemporal changes of metabolites between insects and plants, and discusses its shortfalls and perspectives. The significance of these concerns is beneficial for future insect research such as physiology and metabolism.     

Investigation of protein induction in tumour vascular targeted strategies by MALDI MSI

Characterising the protein signatures in tumours following vascular-targeted therapy will help determine both treatment response and resistance mechanisms. Here, mass spectrometry imaging and MS/MS with and without ion mobility separation have been used for this purpose in a mouse fibrosarcoma model following treatment with the tubulin-binding tumour vascular disrupting agent, combretastatin A-4-phosphate (CA-4-P). Characterisation of peptides after in situ tissue tryptic digestion was carried out using Matrix-Assisted Laser Desorption/Ionisation-Mass Spectrometry (MALDI-MS) and Matrix-Assisted Laser Desorption/Ionisation-Ion Mobility Separation-Mass Spectrometry Imaging (MALDI IMS-MSI) to observe the spatial distribution of peptides. Matrix-Assisted Laser Desorption/Ionisation-Ion Mobility Separation-Tandem Mass Spectrometry (MALDI-IMS-MS/MS) of peaks was performed to elucidate any pharmacological responses and potential biomarkers. By taking tumour samples at a number of time points after treatment gross changes in the tissue were indicated by changes in the signal levels of certain peptides. These were identified as arising from haemoglobin and indicated the disruption of the tumour vasculature. It was hoped that the use of PCA-DA would reveal more subtle changes taking place in the tumour samples however these are masked by the dominance of the changes in the haemoglobin signals.     

Intact protein mass spectrometry and top-down proteomics

American Society for Mass Spectrometry Sanibel meeting on top-down mass spectrometry

St Pete Beach, FL, USA, 24–27 January 2013

Top-down mass spectrometry involves analysis of intact proteins, typically using electrospray ionization, as multiple charging enhances dissociation and thus identification by comparison of precursor and product ion masses with protein sequence databases. Traditionally a low-throughput, precision technology performed on high-resolution Fourier-transform ion cyclotron resonance mass analyzers, top-down proteomics aims to increase throughput for whole proteome analysis while preserving the inherent value of an intact protein mass measurement. This years’ American Society for Mass Spectrometry Sanibel meeting brought together established scientists who have demonstrated the viability of the top-down approach and its applicability to virtually all segments of the proteome, mixing them with researchers from diverse areas and with the common interest of advancing top-down into the high-throughput proteomics mainstream. Advances in instrumentation including the orbitrap analyzer, ionization mechanisms, dissociation strategies and informatics, as well as a wide variety of applications, were discussed in depth, leading to the inescapable conclusion that the future for top-down is bright.     

Molecular mass spectrometry imaging in biomedical and life science research

This review describes the current state of mass spectrometry imaging (MSI) in life sciences. A brief overview of mass spectrometry principles is presented followed by a thorough introduction to the MSI workflows, principles and areas of application. Three major desorption-ionization techniques used in MSI, namely, secondary ion mass spectrometry (SIMS), matrix-assisted laser desorption ionization (MALDI), and desorption electrospray ionization (DESI) are described, and biomedical and life science imaging applications of each ionization technique are reviewed. A separate section is devoted to data handling and current challenges and future perspectives are briefly discussed at the end.     

Qualitative and quantitative mass spectrometry imaging of drugs and metabolites in tissue at therapeutic levels

Mass spectrometry imaging (MSI) is a rapidly evolving technology that yields qualitative and quantitative distribution maps of small pharmaceutical-active molecules and their metabolites in tissue sections in situ. The simplicity, high sensitivity and ability to provide comprehensive spatial distribution maps of different classes of biomolecules make MSI a valuable tool to complement histopathology for diagnostics and biomarker discovery. In this review, qualitative and quantitative MSI of drugs and metabolites in tissue at therapeutic levels are discussed and the impact of this technique in drug discovery and clinical research is highlighted.     

The potential of mass spectrometry imaging in plant metabolomics: a review

Phytochemistry Reviews - Imaging mass spectrometry (MSI) has contributed important information to several scientific fields and more recently to the biological sciences. The use of MSI as an...     

Following the biochemical and morphological changes of Bacillus atrophaeus cells during the sporulation process using Bioaerosol Mass Spectrometry

Bioaerosol Mass Spectrometry (BAMS), a real-time single cell analytical technique, was used to follow the biochemical and morphological changes within a group of Bacillus atrophaeus cells by measuring individual cells during the process of sporulation. A mutant of B. atrophaeus that lacks the ability to produce dipicolinic acid (DPA) was also analyzed. Single cell aerodynamic sizing was used to follow gross morphological changes, and chemical analysis of single cells by mass spectrometry was used to follow some biochemical changes of B. atrophaeus cells during endospore formation.     

Mass processing--an improved technique for protein identification with mass spectrometry data.

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis following tryptic digestion of polyacrylamide gel pieces is a common technique used to identify proteins. This approach is rapid, sensitive, and user friendly, and is becoming widely available to scientists in a variety of biological fields. Here we introduce a simple and effective strategy called "mass processing" where the list of masses generated from a mass spectrometer undergoes two stages of data reduction before identification. Mass processing improves the ability to identify in-gel tryptic-digested proteins by reducing the number of nonsample masses submitted to protein identification database search engines. Our results demonstrate that mass processing improves the statistical score and rank of putative protein identifications, especially with low-quantity samples, thus increasing the ability to confidently identify proteins with mass spectrometry data.     

Mass spectrometry imaging and profiling of single cells

Mass spectrometry imaging and profiling of individual cells and subcellular structures provide unique analytical capabilities for biological and biomedical research, including determination of the biochemical heterogeneity of cellular populations and intracellular localization of pharmaceuticals. Two mass spectrometry technologies-secondary ion mass spectrometry (SIMS) and matrix assisted laser desorption/ionization mass spectrometry (MALDI MS)-are most often used in micro-bioanalytical investigations. Recent advances in ion probe technologies have increased the dynamic range and sensitivity of analyte detection by SIMS, allowing two- and three-dimensional localization of analytes in a variety of cells. SIMS operating in the mass spectrometry imaging (MSI) mode can routinely reach spatial resolutions at the submicron level; therefore, it is frequently used in studies of the chemical composition of subcellular structures. MALDI MS offers a large mass range and high sensitivity of analyte detection. It has been successfully applied in a variety of single-cell and organelle profiling studies. Innovative instrumentation such as scanning microprobe MALDI and mass microscope spectrometers enables new subcellular MSI measurements. Other approaches for MS-based chemical imaging and profiling include those based on near-field laser ablation and inductively-coupled plasma MS analysis, which offer complementary capabilities for subcellular chemical imaging and profiling.     

主元余像集主成分分析在蛋白质质谱数据中的应用

癌蛋白质谱数据中包含了大量未知的内部结构和变量。针对癌蛋白质谱数据这些特点,在总结主元余像集主成分分析（二次主成分分析）应用的基础上,提出了用t-验证方法进行特征子集选取,然后用主元余像集主成分分析提取特征,以线性判别分析进行分类的新方法。通过对典型癌蛋白质谱数据的分类实验,证明该方法不但识别率高,而且需要选取的特征子集小,分类速度快,提高了方法的准确性与分类速度。     

Antidepressant treatment effects on hippocampal protein turnover: Molecular and spatial insights from mass spectrometry

A major challenge in managing depression is that antidepressant drugs take a long time to exert their therapeutic effects. For the development of faster-acting treatments, it is crucial to get an improved understanding of the molecular mechanisms underlying antidepressant mode of action. Here, we used a novel mass spectrometry-based workflow to investigate how antidepressant treatment affects brain protein turnover at single-cell and subcellular resolution. We combined stable isotope metabolic labeling, quantitative Tandem Mass Spectrometry (qTMS) and Multi-isotope Imaging Mass Spectrometry (MIMS) to simultaneously quantify and image protein synthesis and turnover in hippocampi of mice treated with the antidepressant paroxetine. We identified changes in turnover of individual hippocampal proteins that reveal altered metabolism-mitochondrial processes and report on subregion-specific turnover patterns upon paroxetine treatment. This workflow can be used to investigate brain protein turnover changes in vivo upon pharmacological interventions at a resolution and precision that has not been possible with other methods to date. Our results reveal acute paroxetine effects on brain protein turnover and shed light on antidepressant mode of action.     

生物质谱技术及其应用

质谱是带电粒子按质荷比大小顺序排列的图谱,最初主要用来测定元素或同位素的原子量,随着科学的发展及高性能质谱仪器的出现,质谱被越来越多地应用生命科学研究的许多领域,以其质辅助激光解吸附飞行时间质谱和电喷雾质谱为代表的现代生物质谱技术,为蛋白质等生物大分子的研究提供了必要的技术手段。本文在简介近年来比较常用的几种生物质谱技术的基础上,概述了生物质谱技术在蛋白质,核酸研究及检测分析等几个方面的初步应用。     

微重力诱变大肠杆菌快速生长突变株特性

利用旋转培养装置处理大肠杆菌,筛选生长曲线发生变化、提前进入对数期的突变菌株,对菌株进行基因芯片的表达谱分析和质谱分析,研究微重力条件下微生物的生理代谢变化和对微重力条件的适应机制。结果发现突变菌株有114个差异表达基因,其中99个基因表达上调。表达上调基因主要集中在ABC转运系统、糖代谢、三羧酸代谢、磷酸转移酶系统、核酸代谢、脂类代谢等方面。质谱分析从蛋白水平上验证了这个结果。表明经过微重力处理可以筛选到生长加快的菌株,生长加快是菌株相关代谢水平上调的结果。空间微重力通过对微生物生长代谢相关基因的影响来使菌株适应空间环境。     

Emerging technologies in mass spectrometry imaging

Mass spectrometry imaging (MSI) as an analytical tool for bio-molecular and bio-medical research targets accurate compound localization and identification. In terms of dedicated instrumentation, this translates into the demand for more detail in the image dimension (spatial resolution) and in the spectral dimension (mass resolution and accuracy), preferably combined in one instrument. At the same time, large area biological tissue samples require fast acquisition schemes, instrument automation and a robust data infrastructure. This review discusses the analytical capabilities of an "ideal" MSI instrument for bio-molecular and bio-medical molecular imaging. The analytical attributes of such an ideal system are contrasted with technological and methodological challenges in MSI. In particular, innovative instrumentation for high spatial resolution imaging in combination with high sample throughput is discussed. Detector technology that targets various shortcomings of conventional imaging detector systems is highlighted. The benefits of accurate mass analysis, high mass resolving power, additional separation strategies and multimodal three-dimensional data reconstruction algorithms are discussed to provide the reader with an insight in the current technological advances and the potential of MSI for bio-medical research.     

The analysis of underivatized oligosaccharides by matrix-assisted laser desorption mass spectrometry.

Matrix Assisted Laser Desorption Mass Spectrometry is shown to provide a rapid and sensitive technique for the analysis of underivatized oligosaccharides. Typical sample loading is 1 pmol and analysis time is around 5 minutes. Through the use of an internal standard, mass measurements are generally accurate to within 0.5 Da. The technique is particularly useful for the analysis of oligosaccharide mixtures released from glycoproteins.     

Monitoring indole alkaloid production by Penicillium digitatum during infection process in citrus by Mass Spectrometry Imaging and molecular networking

Green mold, caused by Penicillium digitatum, is the most destructive post-harvest disease in citrus. Secondary metabolites produced by fungal phytopathogens have been associated with toxicity to their respective host through the interaction with a wide range of cell targets. Natural products have also been described as important molecules for biocontrol and competition in their respective environment. For P. digitatum, the production of indole alkaloids, tryptoquialanines A and B, have been reported. However, their biological role remains unknown. Mass Spectrometry Imaging (MSI) technique was applied here for the first time to monitor the secondary metabolites produced on the orange surface during infection in order to gain insights about the P. digitatum-citrus interaction mechanisms. Through the combination of MSI and molecular networking it was possible to report, for the first time, the production of tryptoquivalines and fumiquinazolines by P. digitatum and also the accumulation of tryptoquialanines on the fruit surface from 4 to 7 d post inoculation. P. digitatum was also evaluated concerning the ability to sinthesize indole alkaloids in vivo in the different citrus hosts. The biological role of tryptoquialanines was investigated and tryptoquialanine A was submitted to insecticidal bioassays that revealed its high toxicity against Aedes Aegypti, suggesting an important insecticidal action during orange decay.     

微生物学研究中的软电离质谱技术

包括基质辅助激光解吸电离（ＭＡＬＤＩ）和电喷雾（ＥＳＩ）在内的软电离质谱是最近发展起来的质谱技术,由于这些电离方式对样品的破坏性小,质量测定范围大,分子量测定准确,样品纯度要求不高很适合分析成分复杂的微生物样品,ＭＡＬＩＤＩ－ＴＯＦ－ＭＳ结合高分辨率的二维ＳＤＳ－ＰＡＧＥ可以分析１０＾－１２摩尔水平的蛋白,是细菌蛋白质研究过程中必不可少的工具。最近的研究工作表明,通过ＭＡＩＤＩ－ＴＯＦ－ＭＳ或ＨＰ     

Micrometric molecular histology of lipids by mass spectrometry imaging

Time-Of-Flight Secondary Ion Mass Spectrometry is compared to other mass spectrometry imaging techniques, and recent improvements of the experimental methods, driven by biological and biomedical applications, are described and discussed. This review shows that this method that can be considered as a micrometric molecular histology is particularly efficient for obtaining images of various lipid species at the surface of a tissue sample, without sample preparation, and with a routine spatial resolution of 1μm or less.