Raman Microscopy

Confocal Raman Microscopy has many advantages over traditional Raman spectroscopy. For instance, in microscopy small areas of samples can be selected for data collection. The most prominent advantage, however, is the ability to obtain molecular information at different depths of transparent samples. Because of the confocal design of the microscope, any signal originating from the sample that is not in the focal plane of the microscope will not reach the detector.

The WITec 300RA Confocal Raman Microscope (CRM) is used to obtain chemical and structural molecular information in the form of area maps or single point scans. The CRM is widely used for characterization purposes in the following industries:

• Pharmaceutics
• Medical Device
• Semiconductors
• Food and Beverage
• Geology
• Polymers
• Chemical Analysis
• Packaging
• Failure Analysis
• Art and Conservation
• Forensics
• Paints and Adhesives

Typical Experimental Results

Of all the types of imaging modes available on the 300RA CRM, perhaps the two most ubiquitous are depth scanning and chemical mapping. Typical data sets for each are shown below.

Depth Scans

Depth scans are essentially vertical cross sections of samples. The samples do not need to be prepared or microtomed in any particular manner, and often they can simply be placed directly onto the sample stage. The operator can then specify to what depth he or she wants to image. A typical depth scan is shown below. Layer thickness, layer uniformity, and layer identification can all be determined by these depth scans.

Depth scan (left) and the corresponding spectrum of each area (right).

Chemical Maps

The 300RA CRM obtains chemical maps of surfaces by obtaining thousands of rapid point scans and then rasterizing all of the scans to generate an image. The user can then group spectra together using multivariate analyses, filter masks, or by creating basis spectra. The end result is a false-color composite image that indicates the 2D arrangement of the components. A typical area map is shown below.

Chemical maps of three different areas of a sample. Each color corresponds to a distinct component of the sample.

Applications

For more information please read our application notes:

Infrared/Raman/AFM Tri-modal Imaging, PDF

Monitoring Polymer Orientation using Raman Microscopy

2D Chemical Mapping using Confocal Raman Spectroscopy

Instrument: WITec 300RA Confocal Raman Microscope (CRM)

Instrument Key Specifications

Infrared/Raman/AFM Tri-modal Imaging

Multimodal imaging techniques are critical to perform synergistic data acquisition methods. The WITec 300RA Confocal Raman/AFM Microscope is a great illustration of multimodal imaging systems because with a simple rotation of the microscope turret, confocal optical, Raman and AFM imaging can be performed. Furthermore, due to the ease of sample transfer between Ebatco’s Cary Agilent 670 FTIR Microscope, tri-modal imaging can be regularly practiced with little-to-no sample contamination or perturbation. In this application note, a polymer laminate was placed in an FTIR microvice holder and characterized using FTIR spectroscopy first. The microvice sample holder was then lifted in its entirety (without disturbing the sample) and placed under the 20x objective of the confocal Raman/AFM microscope, where the sample was sequentially characterized via Raman and AFM.

In Figure 1, three overlays are provided to illustrate how the confocal Raman-AFM multiple function microscope and FTIR microscope can be used to very easily image a plastic laminate. Because each technique is responsive to different stimuli, unique information is extracted from each type of analysis.

Figure 1. An optical image of a multilayer laminate cross-section taken using the 4x FTIR objective overlaid with an infrared image, an AFM phase image, and a Raman image.

From the top of the image down, the IR overlay is shown first. IR imaging is sensitive to chemical bonds present in each layer, and as such three unique polymers were identified (represented by three colors). The three polymers were found to be layered in such a way to create a six-layered cross-section.

In comparison to IR imaging, AFM phase imaging is not only sensitive to chemical structures, but also other physical and mechanical properties such as elasticity, viscosity, adhesion, and friction. Changes in these surface properties of a polymer can be a result of chemical modification, polymer orientation, or polymer crystallinity, to name a few, all of which affect the resulting contrast in an AFM phase image. Because of this wide variety of effectors, one can isolate over fifteen distinct layers in the AFM image. As observed, changes in surface properties do not necessarily result in spectroscopic changes in the IR overlay.

Finally, Raman imaging is sensitive to chemical bonds present in the sample. However, the selection rules for chemical bonds that can undergo Raman scattering are different than those for that can undergo IR excitation. As such, different bands are present in the Raman spectra than are present in the IR spectra even though the polymers present are identical. Furthermore, the Raman scattering linewidths are markedly smaller than corresponding infrared linewidths, resulting in a more resolved spectrum. It is because of the more resolved Raman spectrum that more information can be obtained. The Raman image in Figure 1 clearly shows at least fourteen bands that are reflective of different chemical bonds, molecular orientations and degrees of crystallinity present in the polymers of the plastic laminate.

Some correlation is certainly observed between the AFM and Raman overlays, indicating the changes in surface properties of the polymers result from the crystallinity or amorphous morphology of the polymer. However, there is also some discrepancy between the images, suggesting that there are other Raman-insensitive factors at play. The AFM bands that do not have corresponding Raman bands could be explained by changes in other surface properties.

As illustrated, with the current instrument suite and setup at Ebatco, samples can be easily transferred between the two imaging microscopes with minimal interruption to perform Infrared/Raman/AFM tri-modal imaging. The multimodal imaging approach helps correlate changes in surface properties to changes in polymer chemical structure, crystallinity and orientation.