SPECTROMETRY & IMAGING
I. Mass Spectrometry Imaging
II. Microspectroscopy - Raman & Brillouin
III. X-Ray Fluorescence Imaging
IV. Energy-Dispersive X-Ray Spectroscopy (EDX)
V. Electron Energy Loss Spectrometry (EELS)
I. Mass Spectrometry Imaging
Mass Spectrometry Imaging (MSI) allows a completely untargeted approach towards tissue visualization. Multiple analytes (drugs, metabolites, proteins, lipids …) are recorded in parallel on tissue. MSI has high potential to be the first method to accomplish the unbiased identification of unknown components involved in biological interactions and to identify compartmentalized biological changes.
Resources
- Cryostat CryoStar NX50 with steel knife and tungsten carbide knife (16 cm each) in addition to standard knifes
- Sample preparation for MS based imaging (sublimation unit, matrix sprayer, picovolume spotter)
- (Atmospheric Pressure) Matrix assisted laser desorption/ionization, (AP-)MALDI
- Desorption Electrospray Ionization, DESI
- Time-of-Flight Mass Spectrometry, ToF-MS for molecular weight determination
- ToF/ToF for MS/MS for analyte identification
- Ion Mobility Separation, IMS
- Software for data interpretation (incl. statistics) and visualization
- Software for multimodal image alignment
Available instrumentation:
- MALDI-ToF/RToF MS w/ MS/MS capability for maximum structural information (CID @20keV)
- MALDI-ToF/RToF MS w/ MS/MS capability for high lateral resolution (low µm)
- AP-MALDI-qToF w/ MS/MS and/or IMS capability for gas phase separation and high mass accuracy (2-5 ppm)
- DESI-qToF w/ MS/MS and/or IMS capability for gas phase separation and high mass accuracy (2-5 ppm)
Applications
- Imaging molecular information on polymer material (e.g explants)
- Imaging of polymer degradation products in tissue
- Imaging of small molecule distributions (drugs, lipids, metabolites, ..) in tissue
- Imaging of peptide/protein/glycoconjugate distributions in tissue
- Imaging of peptides/lipids in cells
II. Microspectroscopy - Raman & Brillouin
A versatile confocal sample scanning imaging spectroscopy setup, built on an inverted microscope, that allows for label free spatial mapping of the Brillouin back-scattering spectra – to determine viscoelastic moduli, Raman scattering spectra, and/or fluorescence/luminescence spectra. In all cases excitation equals 532nm, the spatial resolution is about 1 micron, and the scanning area < 300 microns (larger possible with stitching).
Resources
- Microscope
- Wet lab for sample prep.
- Data analysis workstation & custom software
Applications
- Plant biology
- Cell biology
- Structural biology
III. X-Ray Fluorescence Imaging
Micro-X-Ray Fluorescence Imaging (XRF) performs a qualitative analysis of solid samples. Spatially resolved measurements in air or vacuum with confocal (3D) and non-confocal (2D) setup result in elemental maps. Rh (20W) or Mo (50W) X-ray tubes are used to optimize excitation of low-, medium- and high-Z elements. The fluorescence signal is detected using Si(Li) detectors with the detectable elements ranging from U down to Mg or Na. Scans can be performed with spatial resolution of about 50µm to obtain 2/3D images, nondestructively.
Resources
- Micro-XRF setup
- Sample excitation with two different interchangeable X-ray tubes
- Light microscope to identify the areas of interest
- Confocal and non-confocal setup
- Total reflection X-ray fluorescence
- Sample preparation
- Internal standards
- Vacuum conditions possible
- Measurements in various environmental conditions (air/helium)
- Spectra deconvolution software
- Elemental mapping software
- Quantification software
Applications
- Any kinds of solid samples with flat surface, e.g. tissue cuts, pellets, filters
- Liquid samples (e.g. biological, environmental)
- Particulate matter samples (e.g. aerosols)
IV. Energy-Dispersive X-Ray Spectroscopy (EDX)
Energy Dispersive X-ray spectroscopy (EDX–sometimes also called EDS or EDXS) is a robust and commonly used technique for chemical characterization and imaging. Whereas EDX is fully established for chemical characterization of solid-state samples it is gaining more and more attention for biological materials as well. Biological materials consist mainly of low Z elements. Understanding their structure and composition is of vital interest in biomedical research. For a long time energy dispersive X-ray analysis could not be applied efficiently to such materials because of the low X-ray fluorescence yield for light elements. New detectors show excellent sensitivity and resolution at low X-ray energies. Modern transmission electron microscopes allow chemical characterization of thin samples by EDX with high spatial resolution and high count rates within a short time. Due to their improved sensitivity for low energy X-rays, EDX is also a perfect tool for chemical analysis in scanning electron microscopes allowing the operation at low acceleration voltage and keeping the analytical volume small.
V. Electron Energy Loss Spectrometry (EELS)
EELS is a dedicated analytical technique for light element analysis in the TEM. It can be used for quantitative chemical characterization and energy-filtered imaging and also allows the detection of magnetic properties by EMCD (Electromagnetic circular dichroism).
Resources
- TECNAI F20 FEGTEM (Gatan Image Filter GIF Tridiem (60 kV – 200 kV)
- TECNAI G220 analytical TEM (Gatan Image Filter GIF 2001 (10 kV – 200 kV)
- GATAN Vulcan Cathodoluminescence spectrometer
- Digital micrograph software suite with CL package
Applications
- High resolution light element analysis
- Generation of energy filtered elemental maps
- High resolution CL investigations of marked biomolecules in combination with other EM techniques