Bringing Diagnostic Optical Technologies Into the Clinic.

Wouldn’t itbegreat tohaveadevice inyourexamination room to perform an “optical biopsy”? The human eye alone can get you just so far in dermatology. Even the finest morphologist often requires an invasive biopsy and histopathologic evaluation to arrive at a conclusive diagnosis. Apart from simple dermoscopy, optical interrogationof theskin isnot routinelyused inour field andhas largelybeenconfined to the experimental laboratory, but recentdevelopmentsmaystimulate interest inbringingdiagnosticoptical technologies into the clinic. In this issue of JAMADermatology, Balu et al1 report interesting results fromasmall pilot studydemonstrating the capabilities of an in vivo imaging system to reliably diagnose basal cell carcinomausingaprocessmultiphotonmicroscopy (MPM). Other in vivo optical imaging devices, such as reflectance confocal microscopy (RCM) and optical coherence tomography (OCT), work by detecting variations in the refractive index of cellularandextracellularstructurestoresolvemicroscopicstructure. Relying on refractive index differences as the sole contrastdiscriminant leads to images that are somewhat challenging to interpret.2 Multiphoton imaging systems, in this case MPM, introduce additional degrees of contrast by taking advantage of endogenous fluorophores in the skin such as keratin, collagen, nicotinamide adenine dinucleotide (NADH), flavinadeninedinucleotide(FAD),elastin,andmelanin.Ascanning laser excites these fluorescent molecules in vivo, and the specificemittedwavelengthsarecollectedandusedtocomposean image. Signals at characteristic emission wavelengths distinguish the different fluorophores in the skin. In addition, the concentrationof2of themolecules thatcanbe imaged,namely, NADH and FAD, reflects intrinsic cellular metabolic activity, whichoften varies betweennormal andneoplastic cells. Using a related techniquecalled fluorescent lifetime imagingmicroscopy(FLIM),whichmeasuresthefluorescentdecayrateofaparticular fluorophore,evenmorebiological informationabout the cells and extracellularmatrix can be captured. Comparedwith RCM,MPMallowsonetoconstructadetailedpictureofnotonly thesizeandshapeofcellsbutalsoof thespecificmolecularcomponentsof thecell andextracellular environment.All of this informationcanbesynthesizedtogenerateahigh-resolution,3-dimensional (3-D) image of the living skin. Compared with RCM,MPM is likewatching television in color instead of black andwhite. There are limitationswith themultiphoton system. First, likeconfocal systems ingeneral, the fieldofview(FOV) is small, on the order of 1 to 2 mm2. The acquisition time is slow, and patients must remain still to avoid motion artifact. Therefore, imaging larger lesions becomes problematic and requires generating overlapping “mosaics” and stitching them together digitally. Also, the current generation of devices can only resolve structures to a depth of, at most, 200 μm. A lot of “action” takesplacebelow thisdepth.Newerdevicespromise larger FOVs, more rapid scanning, and deeper penetration, but time will tell if the critical problems can be overcome to produce a practical and affordable device that rivals a punch biopsy in sensitivity and specificity. A combination of systemsmight be used to overcome the limitations of any single system. For instance, our laboratory has published a study3 using an inexpensive reflectance polarizationsystemthatcan imageupto4cm2of skin in real time. The resolution of this system is much lower than a confocal system—by an order of magnitude—but “abnormal” tissue structure can clearly be detected. Using a “macro” and “micro” system in tandem, one could imagine scanning a larger lesionwithamacroviewer to isolate “abnormalities” anda3-D laser scanning system to “zoom in” and identify the specific abnormalities with submicron resolution. A reasonable personmight ask if we even need an optical imaging system for the skin, given the simplicity and reliabilityofa routineskinbiopsy.While itmaypleasepatients toavoid a small biopsy scar, it seems tome that developing the optical biopsy is not particularly urgent for most dermatology patients. A notable exceptionwould be in patients with numerous, difficult-to-track lesions, suchas thosewithbasal cell nevus syndromeordysplastic nevus syndrome.However, inone fieldofdermatology,namelydermatologicsurgery, I thinkthere is indeed an urgent need for an optical biopsy device. AlthoughMohsmicrographic surgery is an elegant, time-tested procedure to remove tumors, it still relies too heavily on the clinician’s visual intuition to “guess” at the initial presurgical margins of the tumor. Not only dowe operate on tumors that are much larger than they seem to be at first glance, we also operatewhen theremayactuallybeno tumorat all, that is, just residual scar from a biopsy. If a surgeon underestimates the presurgical margin, the patient will requiremultiple excision stages, and if he or she overestimates the preoperative margin, too much normal skin is unnecessarily removed. Errorprone guesswork is something we should strive tominimize. Dermatologic surgeons should demand what every other surgical specialty has developed decades ago, namely, an imaging system to assess the presence of tumor and extent of in vivo tumor spread prior to surgery. The challenges facing development of such a system are the requirements for both submicronresolutionthat rivals theresolutionofstandard light microscopy and rapid acquisition of images over a potentially large surface area. Studies attempting to map presurgiRelated article page 1068 Opinion