Distributed computing strategies for processing of FT-ICR MS imaging datasets for continuous mode data visualization.

High-resolution Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry imaging enables the spatial mapping and identification of biomolecules from complex surfaces. The need for long time-domain transients, and thus large raw file sizes, results in a large amount of raw data ("big data") that must be processed efficiently and rapidly. This can be compounded by large-area imaging and/or high spatial resolution imaging. For FT-ICR, data processing and data reduction must not compromise the high mass resolution afforded by the mass spectrometer. The continuous mode "Mosaic Datacube" approach allows high mass resolution visualization (0.001 Da) of mass spectrometry imaging data, but requires additional processing as compared to feature-based processing. We describe the use of distributed computing for processing of FT-ICR MS imaging datasets with generation of continuous mode Mosaic Datacubes for high mass resolution visualization. An eight-fold improvement in processing time is demonstrated using a Dutch nationally available cloud service.

Absorption mode FTICR mass spectrometry imaging.

Fourier transform ion cyclotron resonance mass spectrometry offers the highest mass resolving power for molecular imaging experiments. This high mass resolving power ensures that closely spaced peaks at the same nominal mass are resolved for proper image generation. Typically higher magnetic fields are used to increase mass resolving power. However, a gain in mass resolving power can also be realized by phase correction of the data for absorption mode display. In addition to mass resolving power, absorption mode offers higher mass accuracy and signal-to-noise ratio over the conventional magnitude mode. Here, we present the first use of absorption mode for Fourier transform ion cyclotron resonance mass spectrometry imaging. The Autophaser algorithm is used to phase correct each spectrum (pixel) in the image, and then, these parameters are used by the Chameleon work-flow based data processing software to generate absorption mode "Datacubes" for image and spectral viewing. Absorption mode reveals new mass and spatial features that are not resolved in magnitude mode and results in improved selected ion image contrast.

Advanced mass calibration and visualization for FT-ICR mass spectrometry imaging.

Mass spectrometry imaging by Fourier transform ion cyclotron resonance (FT-ICR) yields hundreds of unique peaks, many of which cannot be resolved by lower performance mass spectrometers. The high mass accuracy and high mass resolving power allow confident identification of small molecules and lipids directly from biological tissue sections. Here, calibration strategies for FT-ICR MS imaging were investigated. Sub-parts-per-million mass accuracy is demonstrated over an entire tissue section. Ion abundance fluctuations are corrected by addition of total and relative ion abundances for a root-mean-square error of 0.158 ppm on 16,764 peaks. A new approach for visualization of FT-ICR MS imaging data at high resolution is presented. The "Mosaic Datacube" provides a flexible means to visualize the entire mass range at a mass spectral bin width of 0.001 Da. The high resolution Mosaic Datacube resolves spectral features not visible at lower bin widths, while retaining the high mass accuracy from the calibration methods discussed.

A novel workflow control system for Fourier transform ion cyclotron resonance mass spectrometry allows for unique on-the-fly data-dependent decisions.

In this paper a novel workflow-based data acquisition and control system for Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) is presented that facilitates a fast on-the-fly decision-making process for a wide variety of data-dependent experiments. Several new workflow implementations demonstrate the flexibility and benefit of this approach for rapid dynamic experimental design on a chromatographic timescale. The different sequence, evaluation, decision and monitoring modules are described using a selected set of examples. During a tandem liquid chromatography (LC)/FTICR-MS experiment the system is used to dynamically switch between various dissociation techniques such as electron capture dissociation (ECD) and sustained off-resonance irradiation (SORI) depending on the charge state of a tryptic peptide peak. The use of this workflow-based system for imaging FTICR-MS using a desorption electrospray ionization (DESI) source demonstrates the possibility of external control of the workflow by feedback from an imaging sample stage.

A software application for comparing large numbers of high resolution MALDI-FTICR MS spectra demonstrated by searching candidate biomarkers for glioma blood vessel formation.

BACKGROUND: A Java application is presented, which compares large numbers (n > 100) of raw FTICR mass spectra from patients and controls. Two peptide profile matrices can be produced simultaneously, one with occurrences of peptide masses in samples and another with the intensity of common peak masses in all the measured samples, using the peak- and background intensities of the raw data. In latter way, more significantly differentially expressed peptides are found between groups than just using the presence or absence in samples of common peak masses. The software application is tested by searching angiogenesis related proteins in glioma by comparing laser capture micro dissected- and enzymatic by trypsin digested tissue sections. RESULTS: By hierarchical clustering of the presence-absence matrix, it appears that proteins, such as hemoglobin alpha and delta subunit, fibrinogen beta and gamma chain precursor, tubulin specific chaperone A, epidermal fatty acid binding protein, neutrophil gelatinase-associated lipocalin precursor, peptidyl tRNA hydrolase 2 mitochondrial precursor, placenta specific growth hormone, and zinc finger CCHC domain containing protein 13 are significantly different expressed in glioma vessels. The up-regulated proteins in the glioma vessels with respect to the normal vessels determined by the Wilcoxon-Mann-Whitney test on the intensity matrix are vimentin, glial fibrillary acidic protein, serum albumin precursor, annexin A5, alpha cardiac and beta actin, type I cytoskeletal 10 keratin, calcium binding protein p22, and desmin. Peptide masses of calcium binding protein p22, Cdc42 effector protein 3, fibronectin precursor, and myosin-9 are exclusively present in glioma vessels. Some peptide fragments of non-muscular myosin-9 at the C-terminus are strongly up-regulated in the glioma vessels with respect to the normal vessels. CONCLUSION: The less rigorous than in general used commercial propriety software de-isotope algorithm results in more mono-isotopic peptide masses and consequently more proteins. Centroiding of peptide masses takes place by taking the average over more spectra in the profile matrix. Cytoskeleton proteins and proteins involved in the calcium signaling pathway seem to be most up-regulated in glioma vessels. The finding that peptides at the C-terminus of myosin-9 are up-regulated could be ascribed to splicing or fragmentation by proteases.