Fig. 3.

Aberration measurements on human skin biopsies. (a)–(d), blue (410–490nm) and red (500–600nm) endogenous 2PEF signals and SHG signal (390nm, shown in green) recorded at 10μm (a), 35μm (b), 60μm (c) and 105μm (d) below skin surface. The strong aberrations introduced by the stratum corneum are evidenced by the darker pattern in deeper images (white ellipses). Media 1 ^{(11.1MB, AVI)} shows the structure of the sample as a function of depth. Scale bar, 100μm. (e), endogenous fluorescence signal at a depth of 80μm (top), and the corresponding recorded spherical aberration amplitude (middle) and total aberration amplitude (bottom). Aberrations increase as a function of depth ( Media 2 ^{(3.6MB, AVI)} ) and reflect the structure of the superficial layer. The blue areas in the middle images and the black areas in the bottom images correspond to an absence of measurement due to a lack of signal from the sample. Scale bar, 100μm. (f), XZ reslice of the data along the middle of the yellow box in (a). The 2PEF and SHG signals are strongly attenuated under the stratum corneum folds (orange arrow) compared with the surrounding regions. (a)–(d) and (f) share the same colour scale shown below (f). (g) and Media 3 ^{(4.4MB, AVI)} , 3D reconstruction of the boxed area in (a). The excitation cones corresponding to the effective NA used for imaging (0.95) are plotted for the two positions corresponding to the red and orange boxed area in (a). The corresponding images are shown as inset at 80μm deep, demonstrating the correlation between the distortion of the excitation wavefront by the skin ridges and the loss of image resolution. Scale bar, 15μm.