Imaging mass spectrometry is emerging as a powerful tool that has been applied extensively for the localization of proteins, peptides, pharmaceutical compounds, metabolites, and lipids in biological tissues. In this article, a three-dimensional mass spectral imaging (3D MSI) technique was developed to examine distribution patterns of multiple neuropeptide families and lipids in the brain of the crab Cancer borealis. Different matrix/solvent combinations were compared for preferential extraction and detection of neuropeptides and lipids. Combined with morphological information, the distribution of numerous neuropeptides throughout the 3D structure of brain was determined using matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-TOF/TOF MS). Different localization patterns were observed for different neuropeptide families, and isoforms displaying unique distribution patterns that were distinct from the common family distribution trends were also detected. In addition, multiple lipids were identified and mapped from brain tissue slices. any biological processes in the body involve interaction and dynamic spatial redistribution of a broad spectrum of compounds. The functions of various biological compounds in the complex tissue or organism are highly related to their locations in tissue structures. The emerging mass spectral imaging (MSI) approach provides an attractive opportunity to detect and probe the complex molecular content of tissues in an anatomical context. The majority of MSI applications have been focused on two-dimensional distribution of analytes of interest, which can only provide information for a thin section from the specimen. This technique has recently been extended to acquire images of serial sections from single specimen and provide a depth dimension to the dataset, allowing three dimensional (3D) representation of multiple analytes in the target organs [1,2]. The in-depth profiling of 3D MSI provides more comprehensive spatial information for molecules of interest, such as localization of proteins and dynamic distribution and partition of pharmaceutical compounds into various target tissues.The application of mass spectrometry to the field of neuroscience has enabled the discovery and characterization of many neuropeptides and neurohormones [3][4][5][6][7][8]. This is usually achieved by tandem MS analysis of peptides fractionated from crude nerve tissue extract. Alternatively, direct tissue methods, in which the tissue is coated with matrix and probed via matrix-assisted laser desorption/ionization (MALDI) MS analysis, enable the sensitive detection of neuropeptides in single organs [9,10] and even single cells [11,12]. More recently, the use of MSI to map the distribution of neuropeptides has gained increased attention [13][14][15]. MSI reduces the time-consuming steps of sample extraction, purification, and separation, while maintaining the topographical information about molecular distribution [16,17]. MALDI MSI enables the study of a broad mass range...

Mass spectrometric imaging (MSI) is a powerful analytical technique that provides two- and three-dimensional spatial maps of multiple compounds in a single experiment. This technique has been routinely applied to protein, peptide, and lipid molecules with much less research reporting small molecule distributions, especially pharmaceutical drugs. This review’s main focus is to provide readers with an up-to-date description of the substrates and compounds that have been analyzed for drug and metabolite composition using MSI technology. Additionally, ionization techniques, sample preparation, and instrumentation developments are discussed.

The described prototype DPS card works well for normal hematocrit whole blood, but further development is needed for samples of much lower or higher hematocrit.

Isobaric tandem mass tags are an attractive alternative to mass difference tags and label free approaches for quantitative proteomics due to the high degree of multiplexing that can be performed with their implementation. A drawback of tandem mass tags are that the co-isolation and co-fragmentation of labeled peptide precursors can result in chimeric MS/MS spectra that can underestimate the fold-change expression of each peptide. Two methods (QuantMode and MS3) have addressed this concern for ion trap and orbitrap instruments, but there is still a need to solve this problem for quadrupole time-of-flight (Q-TOF) instruments. Ion mobility (IM) separations coupled to Q-TOF instruments have the potential to mitigate MS/MS spectra chimeracy since IM-MS has the ability to separate ions based on charge, m/z, and collision cross section (CCS). This work presents results that showcase the power of IM-MS to improve tandem mass tag peptide quantitation accuracy by resolving co-isolated differently charged and same charged peptides prior to MS/MS fragmentation.

Tissue heat stabilization is a vital component in successful mammalian neuropeptidomic studies. Heat stabilization using focused microwave irradiation, conventional microwave irradiation, boiling, and treatment with the Denator Stabilizor T1 have all proven effective in arresting post-mortem protein degradation. Although research has reported the presence of protein fragments in crustacean hemolymph when protease inhibitors were not added to the sample, the degree to which postmortem protease activity affects neuropeptidomic tissue studies in crustacean species has not been investigated in depth. This work examines the need for Stabilizor T1 or boiling tissue stabilization methods for neuropeptide studies of Callinectes sapidus (blue crab) pericardial organ tissue. Neuropeptides in stabilized and non-stabilized tissue are extracted using acidified methanol or N,N-Dimethylformamide (DMF) and analyzed by MALDI-TOF and nanoLC-ESI-MS/MS platforms. Post-mortem fragments did not significantly affect MALDI analysis in the range m/z 650–1600, but observations in ESI MS/MS experiments suggest that putative post-mortem fragments can mask neuropeptide signal and add spectral complexity to crustacean neuropeptidomic studies. The impact of the added spectral complexity did not dramatically affect the number of detected neuropeptides between stabilized and non-stabilized tissues. However, it is prudent that neuropeptidomic studies of crustacean species include a preliminary experiment using the heat stabilization method to assess the extent of neuropeptide masking by larger, highly charged molecular species.