Ion-to-Image, i2i, a Mass Spectrometry Imaging Data Analysis Platform for Continuous Ionization Techniques.

Mass spectrometry imaging (MSI) techniques generate data that reveal spatial distributions of molecules on a surface with high sensitivity and selectivity. However, processing large volumes of mass spectrometry data into useful ion images is not trivial. Furthermore, data from MSI techniques using continuous ionization sources where data are acquired in line scans require different data handling strategies compared to data collected from pulsed ionization sources where data are acquired in grids. In addition, for continuous ionization sources, the pixel dimensions are influenced by the mass spectrometer duty cycle, which, in turn, can be controlled by the automatic gain control (AGC) for each spectrum (pixel). Currently, there is a lack of data-handling software for MSI data generated with continuous ionization sources and AGC. Here, we present ion-to-image (i2i), which is a MATLAB-based application for MSI data acquired with continuous ionization sources, AGC, high resolution, and one or several scan filters. The source code and a compiled installer are available at https://github.com/LanekoffLab/i2i. The application includes both quantitative, targeted, and nontargeted data processing strategies and enables complex data sets to be processed in minutes. The i2i application has high flexibility for generating, processing, and exporting MSI data both from simple full scans and more complex scan functions interlacing MSn and SIM scan data sets, and we anticipate that it will become a valuable addition to the existing MSI software toolbox.

Quantitative determination of sn-positional phospholipid isomers in MSn using silver cationization.

Glycerophospholipids are one of the fundamental building blocks for life. The acyl chain connectivity to the glycerol backbone constitutes different sn-positional isomers, which have great diversity and importance for biological function. However, to fully realize their impact on function, analytical techniques that can identify and quantify sn-positional isomers in chemically complex biological samples are needed. Here, we utilize silver ion cationization in combination with tandem mass spectrometry (MSn) to identify sn-positional isomers of phosphatidylcholine (PC) species. In particular, a labile carbocation is generated through a neutral loss (NL) of AgH, the dissociation of which provides diagnostic product ions that correspond to acyl chains at the sn-1 or sn-2 position. The method is comparable to currently available methods, has a sensitivity in the nM-µM range, and is compatible with quantitative imaging using mass spectrometry in MS4. The results reveal a large difference in isomer concentrations and the ion images show that the sn-positional isomers PC 18:1_18:0 are homogeneously distributed, whereas PC 18:1_16:0 and PC 20:1_16:0 show distinct localizations to sub-hippocampal structures.

Silver-Doped Nano-DESI MSI for Increased Specificity and Sensitivity of Alkenes.

Nanospray desorption electrospray ionization (nano-DESI) is a technique that can be used for mass spectrometry imaging (MSI) experiments. Due to the separation of sampling and ionization events, the nano-DESI solvent can be doped with additives to alter the specificity and selectivity of the experiment. In this chapter, we describe the addition of silver ions to the nano-DESI solvent for increased specificity and sensitivity of double bond containing analyte molecules for nano-DESI MSI.

Determination of Monounsaturated Fatty Acid Isomers in Biological Systems by Modeling MS3 Product Ion Patterns.

Unsaturated free fatty acids are natively present in biological samples as isomers, where double bonds can be situated on different carbons in the acyl chain. While these isomers can have different actions and impacts on biological systems, they are inherently difficult to identify and differentiate by mass spectrometry alone. To address this challenge, several techniques for derivatization of the double bond or metal cationization at the carboxylic group have yielded diagnostic product ions for the respective isomer in tandem mass spectrometry. However, diagnostic product ions do not necessarily reflect quantitative isomeric ratios since fatty acid isomers have different ionization and fragmentation efficiencies. Here, we introduce a simple and rapid approach to predict the quantitative ratio of isomeric monounsaturated fatty acids. Specifically, empirically derived MS3 product ion patterns from fatty acid silver adducts are modeled using a stepwise linear model. This model is then applied to predict the proportion oleic and vaccenic acid in chemically complex samples at individual concentrations between 0.45 and 5.25 µM, with an average accuracy and precision below 2 and 5 mol %, respectively. We show that by simply including silver ions in the electrospray solvent, isomeric ratios are rapidly predicted in neat standards, rodent plasma, and tissue extract. Furthermore, we use the method to directly map isomeric ratios in tissue sections using nanospray desorption electrospray ionization MS3 imaging without any sample preparation or modification to the instrumental setup. Ultimately, this approach provides a simple and rapid solution to differentiate monounsaturated fatty acids using commonly available commercial mass spectrometers without any instrumental modifications.