Bar graph of the mean phase retardation rate with standard error on the mean (deg/ ␮ m) for the hand, temple, and lower back of each of five volunteers. Each data point was obtained from 15 sepa- rate measurements at each location. 

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... on healthy volunteers, re- cruited under a protocol approved by the Institutional Review Board of Massachusetts General Hospital. Each set of OCT images ͑ both intensity and polarization-sensitive images ͒ was acquired, processed, and displayed in 1 s. All images presented here are 5 mm wide and 1.2 mm in physical depth, using the approximation of a constant tissue refractive index of 1.4. Images are displayed on a logarithmic gray scale over a 55 dB dynamic range. Representative conventional and polarization-sensitive images of skin at the lower back and temple regions are shown in Figs. 1 and 2, respectively. The layered structure of skin com- prising epidermal, e, and dermal, d, layers is evident in both conventional images. Features including a hair follicle ͑ * ͒ are observed in the skin of the lower back, which also exhibits a thicker epidermal layer compared to the skin of the temple. A greater density of sebaceous glands in skin of the facial region leads to a less structured appearance within the papillary dermis compared to the lower back, where a strongly backscattering, fibrous layer is observed. Further differences in the properties of skin at these two locations are revealed by the corresponding polarization-sensitive images ͓ Figs. 1 ͑ b ͒ and 2 ͑ b ͔͒ . At both locations, there is minimal change in the polarization state of light within the epidermal layer, as indicated by the image remaining black within this region. However, birefringence within the dermal layers at these skin sites ap- pears visibly different, as indicated by the contrast of the tran- sition from black to white in each image. Skin of the lower back ͓ Fig. 1 ͑ b ͔͒ demonstrates changes in the polarization state of light within the dermal layer, with the measured phase retardation angle cycling over 360° ͑ black to white to black ͒ . In contrast, the lack of strong banding observed in Fig. 2 ͑ b ͒ , corresponding to skin of the temple, indicates considerably lower birefringence at this location. The information contained within these polarization- sensitive images can be displayed quantitatively by calculat- ing the mean phase retardation angle as a function of depth over the full width of an image. Figures 3 ͑ a ͒ and 3 ͑ b ͒ show both the mean intensity and phase retardation corresponding to Figs. 1 and 2. The depth of the secondary peak in the intensity plots indicates the location of the epidermal–dermal boundary, measured at 117 Ϯ 10 ␮ m for the temple and 155 Ϯ 10 ␮ m at the lower back. The phase retardation graphs for both skin locations demonstrate minimal change in the polarization state of light within this epidermal layer, before the accumulated phase retardation increases as light propa- gates within the dermis. Applying a least-squares fit to the linear portion of these phase retardation plots enables skin birefringence to be quantified in terms of the ͑ double-pass ͒ phase retardation rate and associated fitting error, with values of 0.759 Ϯ 0.004 and 0.237 Ϯ 0.004 deg/ ␮ m measured for the lower back and temple locations shown in Figs. 1 and 2, respectively. The observed offset at the tissue surface is due to variance in the computed Stokes parameters that arises from speckle. This noise can be reduced by averaging the Stokes parameters over distances larger than the coherence length of the source. 29 Figure 4 demonstrates how the phase offset observed in Fig. 3 ͑ a ͒ is reduced by averaging the Stokes parameters over different spatial scales in both axial and lateral directions. Stokes parameters were averaged over 160 ␮ m in the lateral direction and 14.1 ␮ m in depth for the graphs presented in Fig. 3. Figures 1 ͑ b ͒ and 2 ͑ b ͒ demonstrate how tissue birefringence can vary within an image: some regions indicate retardation values around 180° ͑ white ͒ while in neighboring regions the retardation remains low ͑ dark gray ͒ . The intention of this study was to examine anatomical variations in skin birefringence rather than small-scale local variations. Therefore, to assign a single retardation value to the region of tissue im- aged, we determined phase retardation values averaged over the full width of each image. To evaluate the reproducibility of our observations, birefringence measurements were taken at the lower back, temple, and back of the right hand, of a group of five healthy male volunteers, aged between 24 and 35 years. For each volunteer, 15 PS-OCT images were taken at each location, consisting of 3 neighboring groups of 5 scans. Each scan was 5 mm wide with successive scans 1 mm apart, such that each group covered a 5 ϫ 5 mm 2 area. Least- squares fits were applied to each phase retardation plot as demonstrated previously, yielding 15 values of phase retardation rate ͑ deg/ ␮ m ͒ with associated individual fitting errors for each location. Mean phase retardation rates were then calculated for the lower back, temple, and hand for each volunteer. The results of these measurements are shown in Fig. 5, with accompanying error bars indicating the standard error on the mean. The measured phase retardation rates exhibit variation according to the skin location, with the temple region exhib- iting the lowest value for all subjects. Conversely, skin of the lower back provided the highest value for all subjects, with skin of the hand yielding intermediate values. Table 1 provides the mean phase retardation rate, standard deviation, and standard error on the mean for data combined from all volunteers, at each of the three locations investigated. To further illustrate the intrinsic variation of skin birefringence with anatomical location, Fig. 6 presents PS-OCT images at the index fingertip of a volunteer. Ridges that form the fingerprint unique to this volunteer are immediately evident in the conventional OCT image, with eccrine ducts extending to the skin surface through the characteristically thick stratum corneum ͑ sc ͒ common to acral skin of the hands and feet. The horizontal black–white banding observed in earlier polarization-sensitive images of skin on the lower back and temple ͓ Figs. 1 ͑ b ͒ and 2 ͑ b ͔͒ , is replaced by three distinct re- gimes of polarization behavior. Upon propagation through the stratum corneum, the polarization state of light varies ran- domly, and is apparent from the random phase retardation angles mapped between black and white within this region. Upon entering the papillary dermis, light is seen to remain in some arbitrary polarization state, with a degree of correlation apparent over spatial scales on the order of 200 ␮ m. Within the lower dermis, depolarization due to multiple scattering dominates and the polarization-sensitive image again becomes randomized. One possible explanation for randomization of the polarization state of light observed in the stratum corneum is that the flattened, keratinized cells present within this layer each act as individual wave plates with random retardation, although this hypothesis requires further investigation. Figure 7 demonstrates the effect of wound healing on skin birefringence; this image was acquired at the site of mature scar tissue on the arm of a volunteer. The image traverses the boundary between normal skin left and scar tissue right , with the scar region providing a slightly stronger ͑ darker ͒ backscattering signal compared to the normal region in Fig. 7 ͑ a ͒ . Examination of the corresponding polarization-sensitive image reveals multiple black–white bands within the scar region that are absent in the normal ͑ left ͒ side of the image. The appearance of strong multiple banding within both young and old scar tissue has been observed consistently during imaging of scar sites on the skin, which we attribute to the presence of newly formed collagen at these locations, which exhibit in- creased birefringence. The polarization properties of the skin provide a contrast mechanism in addition to backscattered light intensity which may be exploited when observing or imaging the skin. PS- OCT enables changes in the polarization state of light to be visualized and quantified in a depth-resolved manner, both at single points in time and over extended courses of time. We have observed the ability of components of the skin to alter the polarization state of light, and attribute this to the birefringent nature of collagen fibers, which along with elastin fibers, provides the dermal layer with mechanical strength and orga- nization. While the presence of collagen is common to skin at all locations on the body, the size, distribution, and relative concentration of individual fibers are known to vary according to the location. Our observations of variable birefringence with location reported here may be understood by consider- ation of the known collagen distribution at the locations ex- amined. The relatively thick skin of the torso and limbs has a thick dermal layer, made up of large bundles of collagen fibers, which we expect to exhibit significant birefringence. Thinner skin covering the face comprises a network of finer collagen bundles, combined with a particularly high density of eccrine glands, resulting in lower birefringence. Collagen remodeling and repair are known to be a fundamental aspect of the wound healing process, and, as we have seen, skin in regions of scar tissue is particularly birefringent. Collagen birefringence has been previously reported, 30,31 as have dynamic changes in birefringence due to thermal denaturation, 31,32 although other potentially influential factors including age, photodamage, and wound healing response have been poorly categorized in vivo . As we have demonstrated here, variation with anatomical location can be significant, although the measurements appear to be consistent across a limited set of samples. PS-OCT provides noninvasive imaging and quantification of both the skin structure and polarization properties in real time. We anticipate that birefringence measurements in human skin ...