Proceedings Article | 13 July 2007
R. Cicchi, S. Sestini, V. De Giorgi, D. Stambouli, P. Carli, D. Massi, T. Lotti, F. Pavone
KEYWORDS: Skin, Luminescence, Tissues, Microscopy, Second-harmonic generation, Fluorescence lifetime imaging, Tissue optics, Tumors, Melanoma, Collagen
We investigated different kinds of human cutaneous ex-vivo skin samples by combined two photon intrinsic fluorescence
(TPE), second harmonic generation microscopy (SHG), fluorescence lifetime imaging microscopy (FLIM), spectral
lifetime imaging (SLIM), and multispectral two photon emission detection (MTPE). Morphological and spectroscopic
differences were found between healthy and pathological skin samples, including tumors. In particular, we examined
tissue samples from normal and pathological scar tissue (keloid), and skin tumors, including basal cell carcinoma (BCC)
and malignant melanoma (MM). By using combined TPE-SHG microscopy we investigated morphological features of
different skin regions, as BCC, tumor stroma, healthy dermis, fibroblastic proliferation, and keloids. A score, based on
the SHG to autofluorescence aging index of dermis (SAAID), was assigned to characterize each region. We found that
both BCC and surrounding dermis have a negative SAAID value, tumor stroma has a positive SAAID value, whereas
fibroblastic proliferation and keloids have a SAAID value close to the unit. Further comparative analysis of healthy skin
and neoplastic samples was performed using FLIM, SLIM, and MTPE. In particular, BCC showed a blue-shifted
fluorescence emission, a higher fluorescence response at 800 nm excitation wavelength and a slightly longer mean
fluorescence lifetime. MM showed an emission spectrum similar to the corresponding healthy skin emission spectrum,
and a mean fluorescence lifetime distribution shifted towards shorter values. Finally, the use of aminolevulinic acid as a
contrast agent has been demonstrated to increase the constrast in BCC border detection. The results obtained represent
further support for in-vivo non-invasive imaging of diseased skin.
