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Optical contrast is a normalised difference between the intensities of light reflected by sample and surrounding substrate
Optical contrast of layered materials changes in a step-like manner with increasing number of layers, allowing accurate thickness estimate for flakes up to 15 layers thick
Optical contrast measurements work best for materials with strong absorption in the visible range, such as graphene, transition metal mono- and dichalcogenides, deposited onto silicon substrates with 90 or 290 nm dioxide layer, but can be extended to other types of substrates
Optical contrast measurements are most commonly used in a qualitative way during flake search to identify monolayers based on their translucent appearance and low optical contrast. An experience user working with a familiar microscope, substrate, and material can identify monolayers with almost 100% accuracy. For more quantitative measurement, optical contrast can be calculated using an image processing software. Figure above shows another example of a WSe2 flake deposited onto PDMS substrate, as well as optical contrast values extracted from mono- and few-layer regions of eight different samples. Even though the contrast values for each thickness show random fluctuations caused by variation of acquisition parameters and non-uniform illumination, the difference between adjacent thicknesses is sufficiently high to accurately estimate the number of layers in each region. Moreover, this difference (around 0.11 for the samples shown above) is equal to the optical contrast of a monolayer region – this allows you to estimate the expected value for a single layer region even before you find one.
Optical contrast is calculated by measuring the intensity of three colour channel (red, green, and blue) in the digital colour image. It can be done using different software, but one of the most convenient choices is ImageJ, a free and power software for microscope image processing.
Acquire an image of the flake, making sure that it is properly focused and exposed
Save the image as a high-quality file (tiff or jpeg with at least 90% quality)
Open the acquired imaged in ImageJ by using the file menu or by dragging and dropping the file onto ImageJ window
Use circular selection tool to select an area of your flake. Press M to perform intensity measurements.
Use mouse to drag the selection onto the substrate, keeping the minimum distance between the measurement points, and repeat the measurements.
Calculate the optical contrast as C = (Isample – Isubstrate)/ (Isample + Isubstrate)
Repeat the measurements for regions with different thickness to estimate the optical contrast expected for monolayer from the minimal difference between adjacent thicknesses.
Calculate the number of layers in the selected region by dividing its optical contrast by the expected monolayer contrast.
Make sure that white balance is set correctly – use a piece of white paper to set it.
Save you images with sufficiently high quality (tiff, jpeg with >90% quality)
Keep the distance between the measurement spots to a minimum
Make sure none of the colour channel are underexposed or clipping
Different optical systems (e.g. microscope lenses, light source, cameras) will provide different contrast – if you are comparing results of different measurements, make sure they were taken using the same system.
Optical contrast can also be affected by acquisition setting (sensitivity, exposure time, WL intensity)
For increases accuracy, consider calculating optical contrast for individual colour channels
If your camera is noisy, take a reference image and subtract the noise background prior to optical contrast calculations
Use another method (AFM, Raman or PL) to verify measurement results
“Rapid and Reliable Thickness Identification of Two-Dimensional Nanosheets Using Optical Microscopy” by H. Li et al. – Identification of thickness for different layered materials up to 15 layers thick deposited onto SiO2/Si substrate with 90 or 300 nm dioxide layer thickness using optical contrast measurements (ACS Nano, arXiv)
“Viscoelastic Transfer of 2D Material Using PDMS” – our detailed text and video guide to 2D material preparation and transfer made in collaboration with Osilla, which includes thickness identification using optical contrast measurements (Osilla website).
“Broad range thickness identification of hexagonal boron nitride by colors” by Y. Anzai – Identification of hBN thickness using its colour for flakes deposited onto 90 nm SiO2/Si substrates (Applied Physics Express)