9 research outputs found
Infrared Matrix-Assisted Laser Desorption/Ionization Orthogonal-Time-of-Flight Mass Spectrometry Employing a Cooling Stage and Water Ice As a Matrix
Although water ice has been utilized in the past as a
matrix for
infrared matrix-assisted laser desorption/ionization mass spectrometry
(IR-MALDI-MS), it has not found a wider use due to limitations in
the analytical performance and technical demands on the employment
of the necessary cooling stage. Here, we developed a temperature-controlled
sample stage for use with an orthogonal time-of-flight mass spectrometer
(MALDI-o-TOF-MS). The stage utilizes a combination of liquid nitrogen
cooling and counterheating with a Peltier element. It allows adjustment
of the sample temperature between ∼−120 °C and
room temperature. To identify optimal irradiation conditions for IR-MALDI
with the water ice matrix, we first investigated the influence of
excitation wavelength, varied between 2.7 and 3.1 μm, and laser
fluence on the signal intensities of molecular substance P ions. These
data suggest the involvement of transient melting of the ice during
the laser pulse and primary energy deposition into liquid water. As
a consequence, the best analytical performance is obtained at a wavelength
corresponding to the absorption maximum of liquid water of about 2.94
μm. The current data significantly surpass the previously reported
analytical features. The particular softness of the method is, for
example, exemplified by the analysis of noncovalently bound holo-myoglobin
and of ribonuclease B. This is also the first report demonstrating
the analysis of an IgG monoclonal antibody (MW ∼ 150 kDa) from
a water ice matrix. Untypical for MALDI-MS, high charge states of
multiply protonated species were moreover observed for some of the
investigated peptides and even for lacto-<i>N</i>-fucopentaose
II oligosaccharides. Using water ice as matrix is of particular interest
for MALDI MS profiling and imaging applications since matrix-free
spectra are produced. The MS and tandem MS analysis of metabolites
directly from frozen food samples is demonstrated with the example
of a strawberry fruit
Calculation of Membrane Lipid Ratios Using Single-Pixel Time-of-Flight Secondary Ion Mass Spectrometry Analysis
Much evidence suggests that membrane
domains, termed lipid rafts,
which are enriched in sphingomyeline and cholesterol play important
roles in the regulation of physiological and pathophysiological processes.
A label-free quantitative imaging method for lipids is lacking at
present. We report an algorithm which enables us to identify and calculate
the percentages of the ingredients of lipid mixtures from single-pixel
time-of-flight secondary ion mass spectrometry (TOF-SIMS) spectra
in model systems. The algorithm is based on a linear mixing model.
Discriminant analysis is used to reduce the dimension of the data
space. Calculations were separately performed for positive and negative
ion mass spectra. Phosphatidylcholine and sphingomyeline which have
identical headgroups and cannot be easily distinguished from another
by positive ion mass spectra were included in the analysis. The algorithm
outlined may more generally be used to calculate the percentages of
ingredients of mixtures from spectra acquired by quite different methods
such as X-ray photoelectron spectroscopy
Water Ice is a Soft Matrix for the Structural Characterization of Glycosaminoglycans by Infrared Matrix-Assisted Laser Desorption/Ionization
Glycosaminoglycans (GAGs) are a class
of heterogeneous, often highly
sulfated glycans that form linear chains consisting of up to 100 monosaccharide
building blocks and more. GAGs are ubiquitous constituents of connective
tissue, cartilage, and the extracellular matrix, where they have key
functions in many important biological processes. For their characterization
by mass spectrometry (MS) and tandem MS, the high molecular weight
polymers are usually enzymatically digested to oligomers with a low
degree of polymerization (dp), typically disaccharides. However, owing
to their lability elimination of sulfate groups upon desorption/ionization
is often encountered leading to a loss of information on the analyte.
Here, we demonstrate that, in particular, water ice constitutes an
extremely mild matrix for the analysis of highly sulfated GAG disaccharides
by infrared matrix-assisted laser desorption/ionization (IR-MALDI)
mass spectrometry. Depending
on the degree of sulfation, next to the singly charged ionic species
doubly- and even triply charged ions are formed. An unambiguous assignment
of the sulfation sites becomes possible by subjecting sodium adducts
of the GAGs to low-energy collision-induced dissociation tandem MS.
These ionic species exhibit a remarkable stability of the sulfate
substituents, allowing the formation of fragment ions retaining their
sulfation that arise from either cross-ring cleavages or rupture of
the glycosidic bonds, thereby allowing an unambiguous assignment of
the sulfation sites
Application of Laser Postionization Secondary Neutral Mass Spectrometry/Time-of-Flight Secondary Ion Mass Spectrometry in Nanotoxicology: Visualization of Nanosilver in Human Macrophages and Cellular Responses
Silver nanoparticles (SNP) are the subject of worldwide commercialization because of their antimicrobial effects. Yet only little data on their mode of action exist. Further, only few techniques allow for visualization and quantification of unlabeled nanoparticles inside cells. To study SNP of different sizes and coatings within human macrophages, we introduce a novel laser postionization secondary neutral mass spectrometry (Laser-SNMS) approach and prove this method superior to the widely applied confocal Raman and transmission electron microscopy. With time-of-flight secondary ion mass spectrometry (TOF-SIMS) we further demonstrate characteristic fingerprints in the lipid pattern of the cellular membrane indicative of oxidative stress and membrane fluidity changes. Increases of protein carbonyl and heme oxygenase-1 levels in treated cells confirm the presence of oxidative stress biochemically. Intriguingly, affected phagocytosis reveals as highly sensitive end point of SNP-mediated adversity in macrophages. The cellular responses monitored are hierarchically linked, but follow individual kinetics and are partially reversible
Application of Laser Postionization Secondary Neutral Mass Spectrometry/Time-of-Flight Secondary Ion Mass Spectrometry in Nanotoxicology: Visualization of Nanosilver in Human Macrophages and Cellular Responses
Silver nanoparticles (SNP) are the subject of worldwide commercialization because of their antimicrobial effects. Yet only little data on their mode of action exist. Further, only few techniques allow for visualization and quantification of unlabeled nanoparticles inside cells. To study SNP of different sizes and coatings within human macrophages, we introduce a novel laser postionization secondary neutral mass spectrometry (Laser-SNMS) approach and prove this method superior to the widely applied confocal Raman and transmission electron microscopy. With time-of-flight secondary ion mass spectrometry (TOF-SIMS) we further demonstrate characteristic fingerprints in the lipid pattern of the cellular membrane indicative of oxidative stress and membrane fluidity changes. Increases of protein carbonyl and heme oxygenase-1 levels in treated cells confirm the presence of oxidative stress biochemically. Intriguingly, affected phagocytosis reveals as highly sensitive end point of SNP-mediated adversity in macrophages. The cellular responses monitored are hierarchically linked, but follow individual kinetics and are partially reversible
Application of Laser Postionization Secondary Neutral Mass Spectrometry/Time-of-Flight Secondary Ion Mass Spectrometry in Nanotoxicology: Visualization of Nanosilver in Human Macrophages and Cellular Responses
Silver nanoparticles (SNP) are the subject of worldwide commercialization because of their antimicrobial effects. Yet only little data on their mode of action exist. Further, only few techniques allow for visualization and quantification of unlabeled nanoparticles inside cells. To study SNP of different sizes and coatings within human macrophages, we introduce a novel laser postionization secondary neutral mass spectrometry (Laser-SNMS) approach and prove this method superior to the widely applied confocal Raman and transmission electron microscopy. With time-of-flight secondary ion mass spectrometry (TOF-SIMS) we further demonstrate characteristic fingerprints in the lipid pattern of the cellular membrane indicative of oxidative stress and membrane fluidity changes. Increases of protein carbonyl and heme oxygenase-1 levels in treated cells confirm the presence of oxidative stress biochemically. Intriguingly, affected phagocytosis reveals as highly sensitive end point of SNP-mediated adversity in macrophages. The cellular responses monitored are hierarchically linked, but follow individual kinetics and are partially reversible
Application of Laser Postionization Secondary Neutral Mass Spectrometry/Time-of-Flight Secondary Ion Mass Spectrometry in Nanotoxicology: Visualization of Nanosilver in Human Macrophages and Cellular Responses
Silver nanoparticles (SNP) are the subject of worldwide commercialization because of their antimicrobial effects. Yet only little data on their mode of action exist. Further, only few techniques allow for visualization and quantification of unlabeled nanoparticles inside cells. To study SNP of different sizes and coatings within human macrophages, we introduce a novel laser postionization secondary neutral mass spectrometry (Laser-SNMS) approach and prove this method superior to the widely applied confocal Raman and transmission electron microscopy. With time-of-flight secondary ion mass spectrometry (TOF-SIMS) we further demonstrate characteristic fingerprints in the lipid pattern of the cellular membrane indicative of oxidative stress and membrane fluidity changes. Increases of protein carbonyl and heme oxygenase-1 levels in treated cells confirm the presence of oxidative stress biochemically. Intriguingly, affected phagocytosis reveals as highly sensitive end point of SNP-mediated adversity In macrophages. The cellular responses monitored are. hierarchically linked, but follow individual kinetics and are partially reversible