Correlated imaging with C60-SIMS and confocal Raman microscopy: Visualization of cell-scale molecular distributions in bacterial biofilms.

Secondary ion mass spectrometry (SIMS) and confocal Raman microscopy (CRM) are combined to analyze the chemical composition of cultured Pseudomonas aeruginosa biofilms, providing complementary chemical information for multiple analytes within the sample. Precise spatial correlation between SIMS and CRM images is achieved by applying a chemical microdroplet array to the sample surface which is used to navigate the sample, relocate regions of interest, and align image data. CRM is then employed to nondestructively detect broad molecular constituent classes-including proteins, carbohydrates, and, for the first time, quinolone signaling molecules-in Pseudomonas-derived biofilms.

Biomolecular imaging with a C60-SIMS/MALDI dual ion source hybrid mass spectrometer: instrumentation, matrix enhancement, and single cell analysis.

We describe a hybrid MALDI/C(60)-SIMS Q-TOF mass spectrometer and corresponding sample preparation protocols to image intact biomolecules and their fragments in mammalian spinal cord, individual invertebrate neurons, and cultured neuronal networks. A lateral spatial resolution of 10 μm was demonstrated, with further improvement feasible to 1 μm, sufficient to resolve cell outgrowth and interconnections in neuronal networks. The high mass resolution (>13,000 FWHM) and tandem mass spectrometry capability of this hybrid instrument enabled the confident identification of cellular metabolites.

MALDI-guided SIMS: multiscale imaging of metabolites in bacterial biofilms.

Mass spectrometry imaging (MSI) is a versatile tool for visualizing molecular distributions in complex biological specimens, but locating microscopic chemical features of interest can be challenging in samples that lack a well-defined anatomy. To address this issue, we developed a correlated imaging approach that begins with performing matrix-assisted laser desorption/ionization (MALDI) MSI to obtain low-resolution molecular maps of a sample. The resulting maps are then used to direct subsequent microscopic secondary ion mass spectrometry (SIMS) imaging and tandem mass spectrometry (MS/MS) experiments to examine selected chemical regions of interest.

Spatial organization of Pseudomonas aeruginosa biofilms probed by combined matrix-assisted laser desorption ionization mass spectrometry and confocal Raman microscopy.

Bacteria growing as surface attached biofilms differ significantly from planktonic cells in several important traits that are reflected in the spatiotemporal organization of the cells and the extracellular polymeric substances they secrete. The structural and chemical features that define these biofilms are explored here using a combination of matrix-assisted laser desorption ionization mass spectrometry (MALDI MS) and confocal Raman microspectroscopies (CRM) to characterize and compare the composition and distribution of biomolecules found in biofilms and planktonic cells of the bacterium Pseudomonas aeruginosa. Three-day old P.

Correlated imaging--a grand challenge in chemical analysis.

Correlated chemical imaging is an emerging strategy for acquisition of images by combining information from multiplexed measurement platforms to track, visualize, and interpret in situ changes in the structure, organization, and activities of interesting chemical systems, frequently spanning multiple decades in space and time. Acquiring and correlating information from complementary imaging experiments has the potential to expose complex chemical behavior in ways that are simply not available from single methods applied in isolation, thereby greatly amplifying the information gathering power of imaging experiments. However, in order to correlate image information across platforms, a number of issues must be addressed.

Progress toward single cell metabolomics.

The metabolome refers to the entire set of small molecules, or metabolites, within a biological sample. These molecules are involved in many fundamental intracellular functions and reflect the cell's physiological condition. The ability to detect and identify metabolites and determine and monitor their amounts at the single cell level enables an exciting range of studies of biological variation and functional heterogeneity between cells, even within a presumably homogenous cell population.

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.

Stretched tissue mounting for MALDI mass spectrometry imaging.

Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) combines information-rich chemical detection with spatial localization of analytes. For a given instrumental platform and analyte class, the data acquired can represent a compromise between analyte extraction and spatial information. Here, we introduce an improvement to the spatial resolution achievable with MALDI MSI conducted with standard mass spectrometric systems that also reduces analyte migration during matrix application.