# Emerging Optical Techniques for the Diagnosis of Onychomycosis
## 3 Imaging Techniques  
### Optical Coherence Tomography (OCT)  
OCT is a non-invasive technique for real-time cross-sectional imaging of living tissues on the micron scale, based on the interferometric measurement of the echo time delay and the magnitude of a back-scattered low-power infrared laser light beam. Light emitted from the source is divided into two beams which are then combined to produce an interference signal. One beam is directed to the tissue sample and reflected by the internal structure, while the other hits a reference mirror. By adjusting the path length of the reference beam, the signal amplitude can be calculated as a function of the depth in the tissue [30,31]. The ballistic (back-scattered) and near-ballistic (back-reflected) photons give the depth information during image reconstruction [32]. This back-scattered intensity constitutes what is usually called an A-line, depicted in Figure 2a. In order to generate a B-scan or two-dimensional scan (panel (b) of Figure 2), several axial scans (A-lines) are obtained at different transverse locations. Finally, by shifting a series of B-scans, a three-dimensional dataset can be achieved, as depicted in Figure 2c. OCT imaging has the potential to discriminate elusive alterations in the nail unit for early detection and diagnosis of fungal infections. Furthermore, being non-invasive, time series analysis of a single sample can be applied to evaluate therapeutic interventions. OCT scans last only a few seconds and since the instrument sends low power density infrared light to the nail (optical window 700-1500 nm) it causes no discomfort to the patient. Moreover, multiple locations of the same nail plate can be screened in parallel. OCT is currently employed in studies as a clinical diagnostic tool for nail disorders. Due to the high spatial resolution, OCT can detect subtle morphological changes to healthy nail units with a higher sensitivity compared to high-frequency ultrasound, demonstrating a clear separation of the nail bed from the nail plate. OCT images clearly showed the layered structure of the nail, the morphological components of the epidermis and the upper dermis, and the area of the nail matrix below the proximal nail plate, along with accurate measurements of the thickness of the nail plate [34]. Onychomycosis is  
Figure 2: Diagram depicting the formation of a tomographic OCT image of a healthy nail and fingertip. (**a**) A-line or axial scan, (**b**) bidimensional scan or B-scan and (**c**) volumetric scan. Custom-built OCT based on Axsun swept-source laser (Axsun, 1310 Swept Source Engine, MA, USA). Reproduced from [33]. depicted in OCT scans as longitudinal hyper-reflective lines within the nail plate. These linear OCT structures seem to correspond to high scattering \"tunnels\" or small \"cleavage levels\" that penetrate the nail plate from the surface inwards and are surrounded by relatively low scattering areas. The structures in Figure 3 correspond to the presence of fungal elements which, due to their high chitin content, reflect light stronger than the surrounding nail components [35]. In addition, infected nails may show a non-uniform layering under the nail plate, a rather sharp transition from the nail plate to nail bed and a 70% thickening of the infected nail [36]. Furthermore, signal shadows due to the increased surface irregularity with inclined lines and spots of higher reflectance capacity were observed, contrary to healthy nails which appeared as homogeneous band-like structures with varying degrees of signal intensity and thickness and an obvious separation of the nail bed [37]. Besides nail plate infection, the identification of morphological attributes of a dermatophytoma, a feature negatively impacting the efficacy of onychomycosis treatments, was also described with OCT [38]. Dermatopmoma was depicted as a clearly demarcated hyper-reflective homogenous accumulation beneath an inhomogeneous nail plate.