Schweizerische Gruppe für Massenspektrometrie
Gruppo svizzero di spettrometria di massa
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Group for Mass Spectrometry Schweizerische Gruppe für Massenspektrometrie |
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Groupe
suisse de spectrométrie de masse Gruppo svizzero di spettrometria di massa |
The next meeting of the SGMS was held on November 25 / 26, 2004, again at the Hotel Dorint on the Beatenberg ... high above Lake Thun in the Bernese Oberland, with a scenic view of the Swiss Alps!
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MALDI-MS imaging is one of the emerging technologies in the study of biomolecules
at surfaces. It provides a wealth of information on the spatial organization
of biomolecules at surfaces. It has the potential of becoming an important diagnostic
tool in biomedical sciences for the differential analysis of diseased versus
healthy tissue. The characterization of the spatial organisation of macromolecules
that are present in complex systems such as cells or ensembles of cells (tissue)
with mass spectrometry can be realized through two approaches: spatial resolved
ion generation (microprobe) or spatial resolved ion detection (microscope) of
the macromolecule(s) of interest.
The highest resolution microprobe images of surfaces are generally obtained
by scanning tightly focused ion beams over surfaces (i.e. SIMS). This technique
is generally used for the study of semi-conductor surfaces as it provides detailed
information on the distribution of a variety of elements and fragment fingerprints
of organic surface molecules. The addition of an acidic matrix to biological
surfaces (similar to the MALDI sample preparation process) aids in the generation
of intact biological molecules, a technique referred to as ME-SIMS. We will
show how this microprobe approach can be used to generate images of biomolecular
distributions in nervous tissue with subcellular spatial resolution.
The fastest mass spectrometric imaging technique is without doubt the mass microscope
approach. The instrumental features and benefits for the rapid study of spatial
organization of macromolecules on biological surfaces with sub-micron spatial
resolution will be described and discussed. It will be shown to improve the
combination of spatial resolution and speed of analysis. This novel stigmatic
mass spectrometric ion imaging instrument records the spatial distribution of
MALDI generated peptide ions over an area of 150 by 150 micrometer with a spatial
resolution exceeding 1 micrometer in a single laser shot. In this new approach
the MALDI-MS spatial resolution is no longer determined by the spot size or
the wavelength of the desorption beam but by the quality of the ion optics and
the spatial resolution of the 2 D detector. Moreover, the molecular flash-photography
approach also allows the usage of different desorption and ionization techniques
that would not deliver useful images with the microprobe approach.
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In the mid of the 17th century, different scientists in Europe,started to dissect
insects and to prepare slices of plants. One discovered bacteria, cells, sperms
(so called animalculae) and much more. The microscope was the most important
tool of these scientists. But, how important is the microscope today in the
field of science?
In my talk, I will show details of the development of the Lightmicroscope and
different microscopic methods in medicine, biology and chemistry since 350 years.
Especially I will present some archeological problems, solved in my Laboratory
within Novartis.
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Polymer samples constitute complex mixtures of large molecules. The different
molecules vary in size (molecular-weight distribution), chemical composition
(chemical-composition distribution), functional groups and end-groups (functionality-type
distribution), architecture (e.g. degree-of-branching distribution, block-length
distribution), regularity (tacticity distribution). Also, many of the properties
of polymers and materials are affected by these variations in molecular structure.
Unavoidably, the characterization of molecular distributions requires separation
techniques.
For polymers, liquid-phase separations appear the obvious choice. Soluble polymers
can be separated according to size using size-exclusion chromatography, field-flow
fractionation or hydrodynamic chromatography. Interactive liquid chromatography
allows separation of macromolecules according to chemical composition or functionality.
Capillary electrophoresis and related techniques provide options for high-resolution
separations.
When several distributions must be characterized simultaneously, multi-dimensional
separation techniques are required. Comprehensive two-dimensional liquid chromatography
is a very useful tool for separation polymers. However, it also poses challenges
by requiring both high-resolution miniaturized separations (first dimension)
and fast, yet efficient separations in the second dimension. If only because
of these requirements contemporary developments in the field of LC, such as
the use of monolithic columns or very high pressures are highly relevant in
the area.
Perhaps more surprising, but equally relevant have been the developments in
mass spectrometry for the characterization of polymers. The emergence of soft
ionization techniques (matrix-assisted laser-desorption ionization, MALDI, and
electrospray ionization, ESI) in combination with Time-of-Flight MS has created
fantastic possibilities. Yet, for characterizing polymer distributions MS techniques
still have there limitations. Both MALDI and ESI are limited to specific classes
of polymers and, most of all, require samples with relatively narrow distributions.
The narrow fractions obtained from one- and two-dimensional separations are
very suitable for analysis by MS. Inversely, the mass and structural information
obtained by MS is invaluable for calibrating and interpreting separation data.
Thus, liquid-phase separations and MS are harmoniously complementary for the
characterization of polymers.
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Mass spectrometry was always dealing with molecules. In addition, to this
molecular mass spectrometry, about 10 years ago, a new type of mass spectrometry
emerged: supramolecular mass spectrometry, which allowed to measures the mass
of non covalent complexes. Complexes of several molecules attached by specific
interactions can be volatilized without destruction of the specific non covalent
interactions.
It took 30 years research effort to the mass spectrometrists community to develop
the methodology for the characterization of tryptic peptides at attomole level
and the automatic interpretation of fragmentation spectra are. Today, this is
routinely used as the key step for the identification of proteins in proteomic
studies. Huge amounts of data are now collected in proteomic studies allowing
to establish the proteomes of cells, tissues, …
We believe that supramolecular mass spectrometry will play a role in what is
the next step after proteomics; the complexomics, but several years of efforts,
if not several decades, will be necessary to fully develop this technology.
In Supra Molecular Mass Spectrometry, the instrument is modified and tuned so
that specific interactions are maintained during the ionisation/volatilization
process.
In these conditions, answers may be given to the following questions:
1. Is there a specific interaction between a protein and a possible ligand?
2. What is the stoichiometry of a multi-protein complex ?
3. Can we detect a cooperativity in the addition of a cofactor?
4. Is there a cooperative effect in the addition of a cofactor ?
Data will be presented where enzyme/ligand interactions are detected and characterized.
Mass measurement of multi-protein complexes of more than 2 million Daltons were
obtained and will be discussed.
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