Confocal Raman Microscopy
Image courtesy of Vladimir Korolkov Photography
Confocal Raman Microscopy
Raman spectroscopy is a non-invasive optical spectroscopy technique used to determine the chemical identity and state of a sample by its unique vibrational modes. The inelastic scattering of light from the sample is measured, with shifts in the energy of the scattered photons being representative of the sample chemistry.
The Raman microscopes at the nmRC can be used as traditional Raman spectrometers to obtain vibrational spectra, but the most powerful feature of these instruments are their ability to collect information from any array of points on - or in - the sample to rapidly produce false colour images (1D, 2D or 3D), where the brightness, contrast and colour of the image is used to convey the science of the sample.
The confocal set-up of the microscopes gives rise to clearer images (by reducing contributions from out-of-focus background regions) and confers the ability to perform depth-profiling studies. These instruments are ideally suited for materials and pharmaceutical characterisation, tissue imaging and chemical identification.
- Conventional and inverted microscope geometries.
- Multiple excitation wavelengths available.
- Multiple gratings available for standard or high spectral resolution spectroscopy.
- Controllable confocality for standard or high spatial resolution hyperspectral imaging.
- SWIFT™ imaging capability for ultra-fast mapping applications.
- Various imaging and software modules tailored for specific applications.
- Variable temperature stage (-196 to 350 oC).
nmRC Confocal Raman Microscopy instrumentation
Horiba LabRAM HR Raman Microscope
- Conventional upright microscope geometry.
- Excitation wavelengths available: 325, 532, 660 and 785 nm.
- Gratings available: 300, 600, 1200 and 1800 lines/mm (depending on configuration).
- Objectives available: 10x, 40x, 50x and 100x (depending on configuration).
- Ultra-low frequency (ULF) Raman module allowing measurement in the sub-100 cm-¹ region (depending on configuration).
- DuoScan™ imaging system for sub micron to macro-scale mapping.
- EasyNav™ software module for optimal Raman imaging.
- Multiwell software module and holder for high throughput screening using traditional well plates and related regular array sample geoometries.
- Multivariate analysis (MVA) software module for advanced data processing.
- Custom-designed flow cell for correlative 'in-situ' dissolution and elution studies.
- Electrochemical cell for Raman spectroelectrochemistry studies (coming soon).
Horiba XploRA INV Raman Microscope
- Inverted microscope geometry, suitable for 'in-situ' analysis and biological samples.
- Excitation wavelength available: 785nm.
- Gratings available: 600, 1200, 1800 and 2400 lines/mm (depending on configuration).
- Objectives available: 10x, 20x and 100x.
One-dimensional mapping: chemical damage to hair
Hair is susceptible to changes and damage induced by a number of factors, including the application of bleaching and colouring treatments. Using confocal Raman microscopy it is possible to spatially locate differences in the composition of (blonde) hair and thus evaluate the extent of damage induced by such chemical treatments. In this study, we line-mapped cross-sections of treated and untreated hair and using the intensity of diagnostic band at 540 cm-1 associated with S-S cross-linkages determined that whilst bleaching does destabilise the structure of the hair at the surface (cuticle), it does not penetrate significantly into the centre (cortex).
Two-dimensional mapping: mechanical exfoliation of graphite
Graphene has been hailed as the wonder material of the twenty-first century. It can be readily procured by simple exfoliation of bulk graphite using sticky tape with the properties of the resultant material dependant on the number of graphitic layers present. Using confocal Raman microscopy it is possible to spatially locate the presence of mono-, bi-, tri- and tetra-layer graphene using the position and symmetry of the diagnostic band at ~2600 cm-1 (2D band). In this study, we used two-dimensional mapping to probe the layering in this few-layer graphene sample and found it to comprise a mixture of layers, a fact that would have been indeterminable from the optical image alone.
Three-dimensional mapping: phase distribution in chocolate
The size, shape and distribution of fats and sugars in confectionary has a remarkable impact on its textural properties which is of high importance to the modern-day consumer. Using confocal Raman microscopy it is possible to spatially locate the presence of the individual ingredients of chocolate based on their diagnostic chemical signatures. In this study, we used three-dimensional mapping to show that the white chocolate under examination comprised small 10-20 micron-sized domains of sugars dispersed within a continuum of fats and milk.