Spectral similarity: Full-wavelength comparisons of circular dichroism spectra

Abstract

Circular dichroism (CD) measurements help characterize optically active molecules and higher-order biomolecular structures. The harmonization of interlaboratory CD measurements requires calibration. Most CD measurements utilize a single wavelength intensity measurement at a spectral peak to calibrate the intensities of the ultraviolet wavelength range. However, such a single-wavelength calibration is inherently less precise and comprehensive than using the CD instrument's spectrum over the entire measured spectral range. A more thorough and informative calibration can be achieved by remapping the spectrum into what we call a Spectral Similarity Plot. This process allows a quantitative evaluation of the shape congruence between two spectra over the full spectral range. Spectral Similarity Plots are highly sensitive to deviations due to differences in concentration, pathlength, source and detector properties, circular polarization balance, as well as wavelength nonlinearities and shifts. Deviations in the totality of these properties can be quantitated by a linear least-squares fit of the remapped data. The remapping enables protocols to correct spectra toward congruence between two spectra. The Spectra Similarity comparison provides an objective test of the CD instrument quality when, e.g., compared to a carefully calibrated system. Finally, we apply the capabilities of the Spectral Similarity Plot to the observation of interferent and binding species in protein samples.