Impact of Scattering Variation Measured by Line-Field Confocal Optical Coherence Tomography on Fluorescence Measurement by Optical Spectroscopy: A Study on Phantoms and Human Skin Models.

introductionSince skin tissues feature highly inter- and intraindividual variable scattering properties, it is of interest for fluorescence spectroscopy applied to skin cancer diagnostic assistance to be combined with a device able to measure scattering properties of skin tissues in vivo and further correct fluorescence spectra. This study aimed to explore the interest of combining two devices previously used in vivo during clinical trials: line-field confocal optical coherence tomography (LC-OCT) for scattering property estimation and fluorescence spectroscopy for measuring the modification of endogenous fluorescence induced by carcinogenesis.

methodsThis study was performed on liquid phantoms and on commercially available in vitro-grown 3D human skin models. Bulk scattering properties of liquid fluorescent phantoms were estimated separately at 800 nm as a function of intralipid concentration from LC-OCT images using a model based on the modified Beer-Lambert law. These results were then compared with values obtained with double-integrating spheres and collimated transmittance measurements followed by inverse adding-doubling estimation of bulk scattering properties. Changes in the amplitude of the chlorin-e6 fluorescence peak were measured as a function of IL concentration using fluorescence spectroscopy. The results obtained on phantoms were then validated with the in vitro-grown skin model.

resultsMeasurements performed on liquid phantoms showed that LC-OCT overestimates scattering coefficient and anisotropy factor by approximately 20-30% compared to values measured by a method (useable only ex vivo) considered here as the gold standard: double-integrating spheres-based optical bench. Fluorescence spectroscopy was employed to measure changes in chlorin-e6 fluorescence-measured intensity relative to varying intralipid concentration. Optical characterization of human skin models confirmed their similarity with in vivo human skin in terms of morphology and of autofluorescence signals. LC-OCT was used to detect dermal scattering coefficient increase induced by impregnating the artificial skin with a PEG-400/DMSO solution that usually acts as an optical clearing agent. However, the observed effect was opposite to the typically expected decrease in the scattering coefficient, which was likely attributable to specific morphological features of the artificial skin that hindered the clearing process, resulting in only hyperosmotic effect. Spectral measurements supported these findings.

conclusionThese findings underscore the interest of combining both optical methods, LC-OCT imaging and autofluorescence spectroscopy, to assess pathology-related fluorophore and scattering alterations in vivo.