Optical coherence tomography in contact dermatitis and psoriasis

Optical coherence tomography (OCT) is a new noninvasive imaging technique. In this study, it was used for the investigation of contact dermatitis and psoriasis. In these common inflammatory skin diseases the value of OCT for quantification and monitoring of the changes in comparison with other bioengineering methods was evaluated. Repeated measurements were performed in healthy volunteers after experimental induction of irritant contact dermatitis and in patients with psoriasis. In the OCT images, the thickness of the epidermis and the signal attenuation coefficient in the upper dermis were evaluated. The changes were compared with measurements of transepidermal water loss, hydration, skin colour and surface roughness, and with high-frequency ultrasound measurements. In irritant dermatitis and psoriasis, thickening of the epidermis was detected and could be monitored over time. The light scattering in the upper dermis was lower than in healthy skin. This was interpreted to be due to the inflammation and oedema, leading to a less-dense arrangement of the collagen fibres. The changes in the OCT images did not significantly correlate with the changes shown by the other methods. OCT is an interesting tool for investigation of inflammatory skin diseases. It is a simple method for determination of epidermal thickness and therefore provides, in addition to other methods, information on the severity of the disease and on treatment effects.     

In vivo reflectance confocal microscopy detects pigmentary changes in melasma at a cellular level resolution

Please cite this paper as: In vivo reflectance confocal microscopy detects pigmentary changes in melasma at a cellular level resolution. Experimental Dermatology 2010; 19 : e228–e233. Abstract: Melasma is a frequent pigmentary disorder caused by abnormal melanin deposits in the skin. In vivo reflectance confocal microscopy (RCM) is a repetitive imaging tool that provides real‐time images of the skin at nearly histological resolution. As melanin is the strongest endogenous contrast in human skin, pigmentary disorders are the most suitable candidates for RCM examination but RCM features of melasma have never been reported. This study investigates the pilot use of RCM in melasma to provide a set of well‐described morphological criteria with histological correlations. RCM images were acquired from melasma skin and compared to adjacent control skin in 26 patients. Skin biopsies were obtained from eight patients. In the epidermis, RCM showed in all patients a significant increase in hyperrefractile cobblestoning cells. These cells corresponded to hyperpigmented basal keratinocytes in histology. In six patients, dendritic cells corresponding to activated melanocytes were also found in the epidermis. In the dermis, RCM identified in nine patients plump bright cells corresponding to melanophages. Interestingly, for a given patient, the topographic distribution of melanophages in melasma lesions was very heterogeneous. RCM also showed a significant increase in solar elastosis and blood vessels in the dermis. RCM is a non‐invasive technique that detects pigmentary changes in melasma at a cellular level resolution. Therefore, RCM provides an innovative way to classify melasma by pigment changes.     

Applications of optical coherence tomography in dermatology

Histology represents the gold standard for morphological investigation of the skin, though biopsy may alter the original morphology, is non-repeatable on the same site and always requires an iatrogenic trauma. In the past decade, advances in optics, fibre as well as laser technology have enabled the development of a novel non-invasive optical biomedical imaging technique, optical coherence tomography (OCT). The latter is based on a classic optical measurement method known as low-coherence interferometry that enables non-invasive, high resolution, two- or three-dimensional, cross-sectional imaging of microstructural morphology in biological tissue in situ. Using conventional OCT with a lateral resolution of 10-15 microm, the stratum corneum of glabrous skin (palmoplantar), the epidermis and the upper dermis can usually be identified, as well as skin appendages and blood vessels. For example, non-invasive monitoring of cutaneous inflammation, hyperkeratotic conditions and photoadaptive processes is possible by means of OCT. Furthermore, the development of high-output broadband light sources, e.g. femtosecond Ti:sapphire laser, might soon enable ultrahigh image resolutions of about 1 microm in order to investigate skin tissue on the cellular level, which could potentially allow the differentiation between benign and malignant tissues. Beyond a high resolution morphology in OCT images, tissue characterization by additional local physical parameters, such as the scattering coefficient and refractive index may be of great value, in particular in cosmetics and the pharmaceutical industry. Functional OCT imaging based on spectroscopy, tissue birefringence, elastography and Doppler flow reveals further information on tissue properties and represents an important progress of OCT technique in the field of dermatology. Therefore, the advanced versions of OCT technique might not only lead to significant new insights in skin physiology and pathology, but also in diagnosis and therapeutic control of cutaneous disorders with respect to non-invasive diagnosis of conditions and monitoring of disease activity in addition to treatment effects over time.     

Changes of human skin in subepidermal wound healing process

Background/purpose: The wound healing process involves unexplained mechanisms. An aberration in this process is known to cause dermal disorders such as keloid or hypertrophic scars, but the mechanism by which these scars are formed remains to be elucidated. Here we examined the usefulness of a non‐invasive optical imaging device to clarify mechanisms of wound healing and of scar formation. Methods: An 8 mm experimental wound was made in the forearms of six subjects by a suction blister method. To observe chronological changes associated with wound healing, horizontal cross‐sectional images were non‐invasively obtained of the wounded area from the skin surface down to 129 μm below at 21.5 μm intervals using in vivo laser confocal scanning microscopy (LCSM). Results: The wounds were covered with a new epidermis by week 2, at which time the dermal papilla count decreased while the thickness from the skin surface to the apex of the dermal papilla increased. The count and the thickness returned to the initial levels when the wound was healed. In two out of six subjects, fibrous tissues were observed in the upper dermis, whereas in one other subject, melanocyte‐like dendritic cells were observed in the epidermis–dermis border in later phases of wound healing. Conclusion: This non‐invasive method using in vivo LCSM revealed chronological changes in the dermis and epidermis during wound healing. In addition, although a scar was not formed in any of study subjects, this microscopy revealed aspects similar to the fibrous tissue overgrowth or to melanocyte migration, both of which may relate to wound healing. These results indicate the usefulness of this non‐invasive method in studies of wound healing and of scar formation.     

High‐definition optical coherence tomography enables visualization of individual cells in healthy skin: comparison to reflectance confocal microscopy

High‐definition OCT (HD‐OCT) is an innovative technique based on the principle of conventional OCT. Our objective was to test the resolution and image quality of HD‐OCT in comparison with reflectance confocal microscopy (RCM) of healthy skin. Firstly, images have been made of a ultra‐high‐resolution line‐pair phantome with both systems. Secondly, we investigated 21 healthy volunteers of different phototypes with HD‐OCT and RCM on volar forearm and compared the generated images. HD‐OCT displays also differences depending on the skin phototype and anatomical site. The 3‐μm lateral resolution of the HD‐OCT could be confirmed by the phantom analysis. The identification of cells in the epidermis can be made by both techniques. RCM offers the best lateral resolution, and HD‐OCT has the best penetration depth, providing images of individual cells deeper within the dermis. Eccrine ducts and hair shafts with pilosebaceous units can be observed depending on skin site. HD‐OCT provides morphological imaging with sufficient resolution and penetration depth to permit visualization of individual cells at up to 570 μm in depth offering the possibility of additional structural information complementary to that of RCM. HD‐OCT further has the possibility for rapid three‐dimensional imaging.     

Optische Kohärenztomographie für die schnelle Bildgebung,Beurteilung und Überwachung entzündlicher Autoimmunerkrankungen der Haut am Krankenbett?

Optical coherence tomography (OCT) is a non‐invasive, high‐resolution imaging technique with a growing impact in dermatology. The principle of OCT is comparable to that of sonography, except that it uses infrared laser light instead of ultrasound waves. It has been clinically demonstrated that OCT is suitable for discriminating between different types of non‐melanoma skin cancer at an early stage of disease. Optical coherence tomography generates two‐ or three‐dimensional images of up to 2 mm penetration depth, a field of view of 6 mm × 6 mm, and an acquisition time of seconds. The resolution capability of OCT is more than 3 to 100 times higher than that of ultrasound imaging. It is of particular interest that the additional information on vasculature provided by OCT angiography enables the assessment and monitoring of inflammatory skin diseases. The use of OCT to locate exact blister levels was demonstrated for diagnosing autoimmune bullous diseases. It is anticipated that detection of subclinical lesions could indicate a relapse of the disease. In the future, this could enable intervention and early treatment. Furthermore, the development of high‐speed OCT could allow fast scanning and bedside imaging of large body sites.     

Optical coherence tomography imaging of bullous diseases

Background Optical coherence tomography (OCT) is a non‐invasive optical imaging technique with a micrometer resolution that may potentially offer real‐time bedside imaging of sufficient detail to allow for morphological discrimination between different types of bullae. Objective To explore the potential of OCT in bullous skin disorders by looking at a set of patients with skin blisters of known origin and study the OCT images for possible hallmarks of the blistering level. Materials and methods OCT provides cross‐sectional, tomographic images of the skin. A consecutive series of patients were recruited and their lesions imaged by OCT: 3 patients with bullous pemphigoid (BP), 1 patient with extensive bullae following burns, 1 patient with pemphigus, 1 patient with subcorneal pustular dermatosis, and a patient with Dariers disease. The latter two were included due to similarity to pemphigus with respect to the level of defect cell adhesion. Results In OCT images, BP bullae are easily depicted as dark, ovoid to round well‐demarquated areas, and BP bulla morphology is clearly different from the burn blisters and the pemphigus‐like disease with respect to the blistering level. Discussion Differentiation of epidermal and subepidermal blisters is demonstrated using OCT. The variation within pemphigoid lesions and pemphigus‐like diseases is however too subtle to allow for differential diagnosis; this may be ascribed to limited resolution. Enhanced resolution of OCT may overcome this obstacle.     

Optical coherence tomography of the human skin

Background: Optical coherence tomography (OCT) is a new diagnostic method for tissue characterization.Objective: We investigated normal and pathologic structures in human skin in several locations to evaluate the potential application of this technique to dermatology.Methods: Based on the principle of low-coherence interferometry, cross-sectional images of the human skin can be obtained in vivo with a high spatial resolution of about 15 μm. Within a penetration depth of 0.5 to 1.5 mm, structures of the stratum corneum, the living epidermis, and the papillary dermis can be distinguished.Results: Different layers could be detected that were differentiated by induction of experimental blisters and by comparison with corresponding histologic sections. Furthermore, OCT images of several skin diseases and tumors were obtained.Conclusion: OCT is a promising new imaging method for visualization of morphologic changes of superficial layers of the human skin. It may be useful for noninvasive diagnosis of bullous skin diseases, skin tumors, and in vivo investigation of pharmacologic effects. (J Am Acad Dermatol 1997;37:958-63.)     

Real‐time three‐dimensional imaging of epidermal splitting and removal by high‐definition optical coherence tomography

While real‐time 3‐D evaluation of human skin constructs is needed, only 2‐D non‐invasive imaging techniques are available. The aim of this paper is to evaluate the potential of high‐definition optical coherence tomography (HD‐OCT) for real‐time 3‐D assessment of the epidermal splitting and decellularization. Human skin samples were incubated with four different agents: Dispase II, NaCl 1 M, sodium dodecyl sulphate (SDS) and Triton X‐100. Epidermal splitting, dermo‐epidermal junction, acellularity and 3‐D architecture of dermal matrices were evaluated by High‐definition optical coherence tomography before and after incubation. Real‐time 3‐D HD‐OCT assessment was compared with 2‐D en face assessment by reflectance confocal microscopy (RCM). (Immuno) histopathology was used as control. HD‐OCT imaging allowed real‐time 3‐D visualization of the impact of selected agents on epidermal splitting, dermo‐epidermal junction, dermal architecture, vascular spaces and cellularity. RCM has a better resolution (1 μm) than HD‐OCT (3 μm), permitting differentiation of different collagen fibres, but HD‐OCT imaging has deeper penetration (570 μm) than RCM imaging (200 μm). Dispase II and NaCl treatments were found to be equally efficient in the removal of the epidermis from human split‐thickness skin allografts. However, a different epidermal splitting level at the dermo‐epidermal junction could be observed and confirmed by immunolabelling of collagen type IV and type VII. Epidermal splitting occurred at the level of the lamina densa with dispase II and above the lamina densa (in the lamina lucida) with NaCl. The 3‐D architecture of dermal papillae and dermis was more affected by Dispase II on HD‐OCT which corresponded with histopathologic (orcein staining) fragmentation of elastic fibres. With SDS treatment, the epidermal removal was incomplete as remnants of the epidermal basal cell layer remained attached to the basement membrane on the dermis. With Triton X‐100 treatment, the epidermis was not removed. In conclusion, HD‐OCT imaging permits real‐time 3‐D visualization of the impact of selected agents on human skin allografts.     

Millimeter wave reflectivity used for measurement of skin hydration with different moisturizers

Background/aims: A new non‐invasive method for determining the free water content in human skin has been developed. The method analyzes the reflection of millimeter (mm) wavelength electromagnetic waves. The amount of reflection of mm waves depends on an electrical property (namely, the permittivity) of the skin, and this depends upon the free water content of the various skin layers. The aim of the present study was to use the mm wave reflectometry method for determination of free water content in healthy skin treated with different hydrating substances. Methods: Skin lotion, pure water, glycerol, and petroleum jelly (an occlusive substance) were used for hydration of skin. The amount of free water was calculated using the permittivity values of skin layers found from fitting a three layer skin model to measured reflection data. The skin model consisted of (1) the stratum corneum (SC), (2) the viable epidermis plus the dermis, and (3) fat layers. Results: Mm wave reflection was significantly affected by the water content of the thick SC of the palm but not by the very thin SC of the forearm. Treatment of the forearm and palm skin with different hydrating substances produced notable changes of the free water content in the SC, but not in the viable epidermis or dermis. The greatest hydration was produced by pure water and skin lotion, and the lowest by petroleum jelly. However, petroleum jelly produced prolonged retention of water in the SC following its hydration by other moisturizers. The content of free water was found to return to its baseline value after removal of moisturizers in as short a time as 8.3 min. Conclusion: The study shows that mm wave reflectometry can be used as a sensitive technique for the non‐invasive determination of water content in living skin.     

In vivo differentiation of common basal cell carcinoma subtypes by microvascular and structural imaging using dynamic optical coherence tomography

The subtype of basal cell carcinoma (BCC) influences the choice of treatment. Optical coherence tomography (OCT) is a non‐invasive imaging tool, and a recent development of an angiographic version of OCT has extended the application of OCT to image the cutaneous microvasculature (so‐called dynamic OCT, D‐OCT). This study explores D‐OCT's ability to differentiate the common BCC subtypes by microvascular and structural imaging. Eighty‐one patients with 98 BCC lesions, consisting of three subtypes: 27 superficial BCC (sBCC), 55 nodular BCC (nBCC) and 16 infiltrative BCC (iBCC) were D‐OCT scanned at three European dermatology centres. Blinded evaluations of microvascular and structural features were performed, followed by extensive statistical analysis of risk ratio (RR) and multiple correspondence analysis. nBCC lesions displayed most characteristic structural and vascular features. Serpiginous vessels, branching vessels, vessels creating a circumscribed figure and sharply demarcated hyporeflective ovoid structures in the dermis were all associated with a higher risk of the subtype being nBCC. The presence of highly present lines and dark peripheral borders at the margin of ovoid structures was negatively associated with iBCC. Lastly, the finding of hyporeflective ovoid structures protruding from epidermis correlated with sBCC. We identified various microvascular and structural D‐OCT features that may aid non‐invasive identification of BCC subtypes. This would allow clinicians to individualize and optimize BCC treatment as well as aid follow‐up of non‐surgical treatment.     

Optical coherence tomography in dermatology: technical and clinical aspects

Optical coherence tomography (OCT), a fairly new non-invasive optical real-time imaging modality, is an emergent in vivo technique, based on the interference (Michelson interferometry) of infrared radiation and living tissues, that allows high-resolution, 2- or 3-dimensional, cross-sectional visualisation of microstructural morphology of tissues. OCT provides depth-resolved images of tissues with resolution up to a few micrometers and depth up to several millimetres depending on tissue type. The investigations using OCT to assess skin structure in clinical settings started in the past decade and consequently proved that this imaging method is useful in visualizing subsurface structures of normal skin, including the epidermis, dermoepidermal junction, dermis, hair follicles, blood vessels and sweat ducts. An increasing number of papers brought evidence of the utility and the precision of OCT technology, in its different technical variants, in diagnosing and monitoring skin disorders, including malignancies and inflammatory conditions, respectively. The present comprehensive review describes and illustrates technical aspects and clinical applications of OCT methods in dermatology.     

Optical coherence tomography (OCT) of collagen in normal skin and skin fibrosis

Optical coherence tomography (OCT) is a non-invasive imaging modality that is transforming clinical diagnosis in dermatology and other medical fields. OCT provides a cross-sectional evaluation of the epidermis and dermis and allows in vivo imaging of skin collagen. Upregulated collagen content is a key feature of fibrotic skin diseases. These diseases are often managed by the practitioner’s subjective assessment of disease severity and response to therapies. The purpose of this review is to provide an overview of the principles of OCT and present available evidence on the ability of OCT to image skin collagen in vivo for the diagnosis and management of diseases characterized by skin fibrosis. We review OCT studies that characterize the collagen content in normal skin and fibrotic skin diseases including systemic sclerosis and hypertrophic scars secondary to burn, trauma, and other injury. We also highlight several limitations of OCT and suggest enhancements to improve OCT imaging of skin fibrosis. We conclude that OCT imaging has the potential to serve as an objective, non-invasive measure of collagen’s status and disease progression for use in both research trials and clinical practice. The future use of OCT imaging as a quantitative imaging biomarker of fibrosis will help identify fibrosis and facilitate clinical examination in monitoring response to treatment longitudinally without relying on serial biopsies. The use of OCT technology for quantification of fibrosis is in the formative stages and we foresee tremendous growth potential, similar to the ultrasound development paradigm that evolved over the past 30 years.     

Pilot study on reflectance confocal microscopy imaging of lichen planus: a real-time, non-invasive aid for clinical diagnosis

Background Lichen planus (LP) represents a relatively common skin inflammatory entity included in the major group of interface dermatitis. In recent years, reflectance confocal microscopy has demonstrated to be a valuable tool for the ‘in vivo’ characterization of various skin diseases with cellular level resolution. No data are currently available that uses reflectance confocal microscopy to study LP. Observations In this study, we have investigated the clinical and confocal features of five cases of histopathologically proven LP, and we have correlated the observed features with histopathological findings. The most characteristic criterion was the presence of interface dermatitis. Papillary rims, usually visible in normal skin, were obscured by the presence of a diffuse inflammatory cells infiltrate, arranged in sheet‐like structures that surrounded the junction almost completely. There was an almost total obliteration of the ring‐like structures around DP, which appeared non‐edged and non‐rimmed. Granular cells appeared as very large, polygonal structures, with an evident grainy cytoplasm, with the transition between spinous and granular cells being clearly recognizable, and this feature corresponded to hypergranulosis in histology. The presence of inflammatory cells at the level of the epidermis was seen as round‐to‐polygonal bright structures in the context of a variable degree of epidermal disarray and spongiosis. Melanophages in dermis were visible as brightly refractile, plump, oval to stellate‐shaped cells. Prominent round or linear dark canalicular structures corresponded to dilated blood vessels in the superficial dermis on histopathology and appeared horizontally oriented in confocal sections. Conclusions Reflectance confocal microscopy may represent a real‐time, non‐invasive aid to clinical diagnosis of LP. However, it might be difficult to distinguish between different subtypes of interface dermatitis. Further research, including larger case series, will better define a possible differential diagnosis of these diseases using confocal microscopy.     

Acute skin alterations following ultraviolet radiation investigated by optical coherence tomography and histology

Optical coherence tomography (OCT) appears to be a promising technique to study skin in vivo. As part of an exploratory study to investigate UV induced effects non-invasively we aimed to evaluate the kinetics of acute UVB- as well as UVA1 induced skin alterations by means of OCT, and to correlate the results obtained with routine histology. Twelve healthy subjects received daily 60 J/cm² of UVA1 and 1.5 minimal erythema doses of UVB on their upper back over three consecutive days. One day (24 h) after the last UV exposure, OCT measurements and skin biopsies were performed in four subjects (day 1) on the centre of the irradiated sites and an adjacent non-irradiated control site. The same procedure was performed in four subjects 3 days and 6 days after irradiation, respectively. Prior to OCT assessment two waterproof marks were drawn on the centre of UVB and UVA1 exposed sites and the control site. The OCT scanner, SkinDex 300, was used in the RI1D measurement modus in order to investigate morphological features, epidermal thickness, and scattering coefficients. Immediately after OCT assessment, 4 mm punch biopsies were taken from the previously marked sites. OCT as well as histological examinations performed on day 1, 3, and 6, revealed markedly higher values for epidermal thickness on UVB exposed skin sites, and slightly increased epidermal thickening in UVA1 exposed sites. UVB exposed sites showed disruption of the entrance signal in the B-scan of OCT resulting in a thickened layer with a signal-poor centre corresponding to hyperkeratosis and parakeratosis as confirmed by routine histology. Surprisingly, the mean scattering coefficients of the epidermis were slightly lower on UVA1 exposed sites, as compared to non-irradiated skin. By contrast, the scattering coefficient of the upper dermis of UVA1 irradiated skin was hardly altered. Moreover, the scattering coefficient of the upper dermis assessed on UVB exposed skin on day 1 was clearly smaller than the scattering coefficient observed on non-irradiated and UVA1 exposed skin. Conclusively, it was possible to demonstrate by means of OCT differences of epidermal thickness and pathological features of the stratum corneum following UV exposure. UVA1 induced epidermal pigmentation as well as UVB induced dermal inflammation may affect the light attenuation in the tissue indicated by a decrease of the scattering coefficient. OCT seems to be a useful tool to monitor UV induced effects in vivo.     

Combining confocal Raman microscopy and freeze‐drying for quantification of substance penetration into human skin

In the area of dermatological research, the knowledge of rate and extent of substance penetration into the human skin is essential not only for evaluation of therapeutics, but also for risk assessment of chemicals and cosmetic ingredients. Recently, confocal Raman microscopy emerged as a novel analytical technique for analysis of substance skin penetration. In contrast to destructive drug extraction and quantification, the technique is non‐destructive and provides high spatial resolution in three dimensions. However, the generation of time‐resolved concentration depth profiles is restrained by ongoing diffusion of the penetrating substance during analysis. To prevent that, substance diffusion in excised human skin can instantly be stopped at defined time points by freeze‐drying the sample. Thus, combining sample preparation by freeze‐drying with drug quantification by confocal Raman microscopy yields a novel analytical platform for non‐invasive and quantitative in vitro analysis of substance skin penetration. This work presents the first proof‐of‐concept study for non‐invasive quantitative substance depth profiling in freeze‐dried excised human stratum corneum by confocal Raman microscopy.     

Optical coherence tomography: its role in daily dermatological practice

Optical coherence tomography (OCT) is a non‐invasive, tomographic imaging technique which generates high‐resolution in‐vivo images up to mid‐dermal layers. Due to continuous technological improvements, OCT is moving from research projects into daily dermatological practice. It can complement other imaging methods like high‐frequency ultrasound or confocal microscopy. There is a wide variety of indications for OCT. In addition to aiding in the diagnosis and clinical monitoring of inflammatory dermatoses, OCT is a very useful and feasible technique in dermato‐oncology.     

In vivo optical coherence tomography of basal cell carcinoma

BACKGROUND: Optical coherence tomography (OCT) is a promising non-invasive imaging technique that has not systematically been studied in skin cancer such as basal cell carcinoma (BCC). OBJECTIVE: We aimed, first, to describe the in vivo histologic features of BCC by using OCT, and second, to find out whether it is possible to differentiate BCC subtypes by means of OCT. METHODS: Prior to the excision, the BCCs (n=43) as well as adjacent non-lesional skin sites were assessed by OCT in vivo. The lesional area of interest was marked prior to OCT and tattooed after excision, respectively, in order to enable topographical concordance between the cross-sectional OCT images and the histologic sections. RESULTS: Compared to non-lesional skin, a loss of normal skin architecture and disarrangement of the epidermis and upper dermis was observed in the OCT images of BCCs. Features that were frequently identified by OCT and correlated with histology included large plug-like signal-intense structures, honeycomb-like signal-free structures, and prominent signal free cavities in the upper dermis. With regard to the aforementioned OCT features, no statistically significant (P<0.05) difference was found between nodular, multifocal superficial, and infiltrative BCCs, respectively. CONCLUSIONS: OCT is capable to visualize altered skin architecture and histopathological correlates of BCC. However, there is not at this time sufficient data supporting the clinical use of OCT for the differentiation of BCC subtypes.     

Validation of a diagnostic algorithm for the discrimination of actinic keratosis from normal skin and squamous cell carcinoma by means of high‐definition optical coherence tomography

Actinic keratoses (AKs) commonly arise on sun‐damaged skin. Visible lesions are often associated with subclinical lesions on surrounding skin, giving rise to field cancerization. To avoid multiple biopsies to diagnose subclinical/early invasive lesions, there is an increasing interest in non‐invasive diagnostic tools, such as high‐definition optical coherence tomography (HD‐OCT). We previously developed a HD‐OCT‐based diagnostic algorithm for the discrimination of AK from squamous cell carcinoma (SCC) and normal skin. The aim of this study was to test the applicability of HD‐OCT for non‐invasive discrimination of AK from SCC and normal skin using this algorithm. Three‐dimensional (3D) HD‐OCT images of histopathologically proven AKs and SCCs and images of normal skin were collected. All images were shown in a random sequence to three independent observers with different experience in HD‐OCT, blinded to the clinical and histopathological data and with different experience with HD‐OCT. Observers classified each image as AK, SCC or normal skin based on the diagnostic algorithm. A total of 106 (38 AKs, 16 SCCs and 52 normal skin sites) HD‐OCT images from 71 patients were included. Sensitivity and specificity for the most experienced observer were 81.6% and 92.6% for AK diagnosis and 93.8% and 98.9% for SCC diagnosis. A moderate interobserver agreement was demonstrated. HD‐OCT represents a promising technology for the non‐invasive diagnosis of AKs. Thanks to its high potential in discriminating SCC from AK, HD‐OCT could be used as a relevant tool for second‐level examination, increasing diagnostic confidence and sparing patients unnecessary excisions.