The Raman Spectroscopy FAQ
Raman spectroscopy can get technical fast. This FAQ cuts through the jargon and gives you clear, honest answers to some of the most common questions, whether you’re new to Raman or just need a quick fact check.
Raman answers.
Just what you need to know.
Basics
- What are the Raman effect and Raman scattering?
The Raman effect describes the inelastic scattering of light by molecules, where scattered photons change energy in accordance with molecular vibrations. Raman scattering refers specifically to those inelastically scattered photons and forms the basis for identifying molecular structure, crystal form and chemical bonds. - Why is the horizontal axis of the Raman spectrum displayed in wavenumber?
Although Raman-scattered light contains a range of wavelengths, Raman spectra are typically displayed in wavenumber rather than wavelength because Raman scattering originates from changes in intrinsic molecular vibrational energy. Since frequency corresponds to the reciprocal of the period (and thus the reciprocal of wavelength), expressing the spectral axis in wavenumber allows the vibrational frequency of the molecule to be determined directly. This makes it possible to estimate the molecular species and also ensures that spectra can be compared regardless of the excitation wavelength. Displaying Raman spectra in wavenumber additionally enables straightforward comparison with infrared absorption spectra. - Is quantitative measurement possible with Raman spectroscopy?
Raman spectroscopy cannot yield absolute quantitative values. However, quantitative analysis becomes possible by preparing a calibration curve using a reference sample. - What sample sizes can be measured?
The maximum sample size is generally limited by the stage capacity to approximately 5 cm × 7 cm × 2.5 cm in thickness. The minimum sample size can be as small as a powder particle, as long as it can be located and brought into focus under the microscope. - What determines the measurement time in Raman imaging?
With line illumination, 400 pixels in the X-direction are acquired with each irradiation. The imaging area is expanded by stepping through positions in the Y-direction, so the measurement time is obtained by multiplying the irradiation time per line by the number of Y-direction pixels. CCD data transfer time must also be considered, and the required irradiation time depends on the Raman activity of the sample and on how sensitive it is to thermal damage. - Can surface morphology be measured with a Raman microscope?
A laser Raman microscope does not provide numerical measurements of height or surface roughness. If the surface roughness is not visually significant in the optical microscope image, it generally will not strongly affect Raman imaging. - What is Surface-Enhanced Raman Scattering (SERS)?
SERS is an amplification phenomenon that occurs when molecules are adsorbed on or near nanostructured metallic surfaces such as gold or silver. The local electromagnetic “hot spots” dramatically increase Raman signal intensity, enabling detection of very low concentrations.
Technical Details
- How do you obtain the best Raman spectral resolution?
Spectral resolution depends on parameters such as the laser wavelength, the groove density of the diffraction grating and especially the spectrometer’s focal length. Higher grating densities and longer focal lengths provide finer spectral resolution. - What is the relationship between spot size, spatial resolution, and pixel size?The laser spot size refers to the diameter of the laser beam focused by the objective lens. In theory, it is governed by the laser wavelength and the numerical aperture of the objective lens, but in practice it also depends on whether the illumination optical system has been properly designed. Spatial resolution is often treated as equivalent to spot size for practical purposes, yet the two differ conceptually because spatial resolution additionally depends on the performance of the detection optics that collect the Raman-scattered light. Micro-Raman instruments usually employ a confocal optical system, and reducing the aperture of the pinhole or slit can, in principle, make the spatial resolution smaller than the spot size. However, such adjustments are not very practical because they significantly reduce the amount of light reaching the detector. Pixel size in Raman imaging refers to the scanning pitch that determines how finely spatial information is sampled. According to the Nyquist theorem, choosing a pixel size approximately half the spatial resolution is sufficient.
- What are sensitivity and brightness in Raman spectroscopy?
Because Raman scattering is inherently weak, high sensitivity and brightness are always desirable. The brightness of a spectrometer is described by its f-number, which becomes smaller and brighter as the focal length decreases. However, the critical factor for optical efficiency is not the absolute f-number but the match between the f-number of the spectrometer and that of the incident optical system. If the incident light is too bright relative to the spectrometer’s f-number, the light spreads beyond the diffraction grating and forms stray light. Conversely, if the spectrometer is brighter than the input optics, the light fails to illuminate the entire grating surface and the optical system becomes underutilized. Commercial instruments vary in f-number, but each design matches internal optical components appropriately; therefore, it is not correct to assume that a spectrometer with a smaller f-number necessarily offers better sensitivity. - Is there a problem with accommodating UV to near-infrared wavelengths in one instrument?
Caution is required when incorporating lasers of widely differing wavelengths into a single instrument. Optical components, especially lenses, have specific operational wavelength ranges, and their performance declines significantly outside those ranges. This is why microscope manufacturers provide different objective lenses for the ultraviolet, visible, and near-infrared regions. The same consideration applies to the internal optical components of a Raman spectrometer. Although some parts may be shared among lasers with similar wavelengths, attempting to use identical optical components across the entire UV to near-infrared range typically leads to insufficient performance in all spectral regions. Instruments designed to be “all-purpose” may end up performing poorly or may ultimately be usable at only one wavelength. It is therefore essential to confirm whether an instrument meets the specifications required for the intended measurement. - How deep does Raman scattering probe into a sample?
The intensity of light decreases exponentially as it passes through a material according to the relation I = I₀e⁻ᵅˣ, where I₀ is the initial intensity, I the transmitted intensity, x the distance traveled, and α the absorption coefficient. The penetration depth L₀, defined as the depth at which the light intensity becomes 1/e of its original value, is therefore 1/α. In Raman scattering, both the incident and the scattered light experience absorption, so the effective sampling depth is roughly half of the standard penetration depth, or 1/(2α). For example, crystalline silicon at 532 nm has an absorption coefficient of approximately 10⁴ cm⁻¹, corresponding to an information depth of about 500 nm. - Does the laser spot distort when scanning near the edge of the field of view?
It is a common misconception that laser beam scanning distorts the spot at the edges of the field of view. In a properly engineered system, the laser beam remains perpendicular to the sample surface whether it is directed at the center or the edge, so the spot shape is maintained across the entire field.
Applications
- How is a Raman image created?
Each pixel in the measurement area contains a full Raman spectrum. When a color is assigned to a peak at a specific wavenumber, the intensity of that peak in each pixel’s spectrum determines the color tone at that pixel and represents the spatial concentration distribution of the associated component. By assigning different colors to peaks associated with different components, it becomes possible to analyze the spatial distribution of multiple materials simultaneously. - How can the cross-sectional structure of plastic wrap be measured non-destructively?
Our Raman microscopes provide a Z-axis spatial resolution of approximately 1 µm thanks to its high-quality confocal optical system. As a result, when dealing with transparent materials, it is possible to focus inside the sample and extract Raman signals specifically from the selected depth. Even multilayer plastic films with layer thicknesses of only a few hundred nanometers can be analyzed non-destructively by adjusting the focal position from the surface inward. - Does surface roughness affect Raman imaging and how can it be handled?
Confocal Raman microscopes are not ideal for highly uneven surfaces because out-of-focus regions are deliberately excluded. If the sample’s surface roughness is modest and does not appear problematic under the optical microscope, Raman imaging is usually unaffected. When roughness causes issues, the impact can be reduced by using a long working distance objective lens of the same magnification. In addition, the 3D-Build function can combine only the in-focus parts of images taken at different heights, thereby improving imaging for samples with significant height variations. For even larger roughness on the order of about 1 mm, a wide-field Raman scope can perform imaging analysis with reduced blurring. - Can you perform in-situ Raman analysis?
Yes. Raman measurements can be performed inside electrochemical cells, reaction chambers or other working environments by using optical windows or fibre-optic probes, enabling real-time monitoring of processes. - Can Raman be used to measure thin-film thickness on transparent substrates?
Raman can measure film thickness on transparent substrates such as glass or polymers, but depth resolution is limited by the optical diffraction limit. Axial resolution is typically around one micron, making sub-micron thickness measurements unreliable. - Can Raman measurements be performed in a controlled environment?
Raman spectroscopy can be used inside controlled environments such as vacuum chambers or isolated sample enclosures, provided an optical window is present to allow both excitation and collection of the scattered light.
Maintenance and Support
- Do lasers have a lifespan?
Yes. Expect to replace the laser after about 5 years due to diode degradation.