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Circular Dichroism Spectral Similarity Plots to Extend Validation and Correction to All Measured Wavelengths



Curtis Meuse


Interlaboratory comparisons of circular dichroism (CD) spectra are useful for developing confidence in the measurements associated with biopharmaceuticals. These measurements help define a drug's higher order (secondary and tertiary) structure. Unfortunately, the extent of the validity of these measurements is unclear. In this work, a method is described to extend CD validation over the entire observed wavelength range using what I call spectral similarity plots. The method involves plotting, wavelength by wavelength, all measured spectral intensities of a sample at one concentration against the intensity values of the same material at a different concentration. These spectral similarity plots validate the instrument in terms of spectral shape and where that shape shifts vertically and/or horizontally. This comparison tests the linearity of instrument's signal, the balance of its left and right polarization as well as its wavelength and spectral intensity scales. When the process is applied to materials with accepted and archived intensity values, the method can be linked to older single- and double-wavelength calibration techniques. Further, spectral similarity testing of CD spectra from samples with different concentrations run in different labs suggests that improved inter-laboratory validation of CD data is possible. Since a database of archival-CD measurements is available online, spectral similarity comparisons could possibly provide the ability to compare linearity, polarization balance, wavelength and spectral intensity between all current CD instruments. If the preliminary results published here prove robust and transferable, then comparisons of full-wavelength-range spectra to archived data using spectral similarity plots should become part of the standard process to validate and calibrate the performance of CD instruments.
Applied Spectroscopy


Circular dichroism, validation, intensity and wavelength linearity, full-wavelength range


Meuse, C. (2023), Circular Dichroism Spectral Similarity Plots to Extend Validation and Correction to All Measured Wavelengths, Applied Spectroscopy, [online],, (Accessed June 15, 2024)


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Created March 1, 2023, Updated March 19, 2023