Basal cell carcinoma characterization using fusionex vivoconfocal microscopy: a promising change in conventional skin histopathology

Abstract

Summary Background Ex vivo confocal microscopy (CM) works under two modes, fluorescence and reflectance, allowing the visualization of different structures. Fluorescence CM (FCM) requires a contrast agent and has been used for the analysis of basal cell carcinomas (BCCs) during Mohs surgery. Conversely, reflectance CM (RCM) is mostly used for in vivo diagnosis of equivocal skin tumours. Recently, a new, faster ex vivo confocal microscope has been developed which simultaneously uses both lasers (fusion mode). Objectives To describe the BCC features identified on reflectance, fluorescence and fusion modes using this novel device. To determine the best mode to identify characteristic BCC features. To develop a new staining protocol to improve the visualization of BCC under the different modes. Methods From September 2016 to June 2017, we prospectively included consecutive BCCs which were excised using Mohs surgery in our department. The lesions were evaluated using ex vivo CM after routine Mohs surgery. The specimens were first stained with acridine orange and then stained using both acetic acid and acridine orange. Results We included 78 BCCs (35 infiltrative, 25 nodular, 12 micronodular, 6 superficial). Most features were better visualized with the fusion mode using the double staining. We also identified new CM ex vivo features, dendritic and plump cells, which have not been reported previously. Conclusions Our results suggest that nuclei characteristics are better visualized in FCM but cytoplasm and surrounding stroma are better visualized in RCM. Thus, the simultaneous evaluation of reflectance and fluorescence seems to be beneficial due to its complementary effect. What's already known about this topic? Ex vivo fluorescent confocal microscopy (FCM) is an imaging technique that allows histopathological analysis of fresh tissue. FCM is faster – at least one‐third of the time – than conventional methods. FCM has a sensitivity of 88% and a specificity of 99% in detecting basal cell carcinomas (BCCs). What does this study add? Reflectance and fluorescence modes can be used simultaneously in a new ex vivo CM device. Each mode complements the other, resulting in an increase in the detection of BCC features in fusion mode. A combined staining using acetic acid and acridine orange enhances the visualization of tumour and stroma without damaging the tissue for further histopathological analysis.

The road to real‐time, bedside, optical imaging pathology: basal cell carcinoma and beyond

“…In this issue of the BJD, P erez-Anker et al report their experience of using FuCM on 78 patients with BCC [60% of them showing a high-risk histological subtype (45% infiltrative, 32% nodular, 15% micronodular, 8% superficial)]. 11 With a mean time of 7 min (range 3-15) required to scan each sample, P erez-Anker et al found that FuCM is beneficial owing to the synergistic effect of visualizing both RCM and FCM features simultaneously, overcoming the individual limitations of both RCM and FCM. Overall, nuclei characteristics were better seen with the FCM and cytoplasm, and surrounding stroma were better seen with RCM.…”

“…similar to polarized and non-polarized dermoscopy complementing each other). 11 The importance of the novel FuCM device is that it improves the visualization of nuclear, cytoplasmic and stromal details by simultaneously obtaining the three CM modes: RCM, FCM and FuCM. Acetic acid and acridine orange double stains may enhance different morphological features of BCC in FuCM and thus potentially aid in cancer margin assessment.…”

“…Also, when compared with the traditional MMS tissue workflow, ex vivo CM can reduce the amount of time required for tissue processing by up to two‐thirds . Recently, a new‐generation ex vivo CM was able to acquire RCM and FCM images simultaneously, to generate ‘fusion’ confocal images (FuCM), in which greyscale RCM and green FCM images are superimposed onto each other. Such images can be even further processed with digital staining, i.e.…”

“…In this issue of the BJD, Pérez‐Anker et al . report their experience of using FuCM on 78 patients with BCC [60% of them showing a high‐risk histological subtype (45% infiltrative, 32% nodular, 15% micronodular, 8% superficial)] . With a mean time of 7 min (range 3–15) required to scan each sample, Pérez‐Anker et al .…”

“…However, the authors highlight that FuCM should not be used by itself, but rather it should be a complement to both FCM and RCM modes owing to specific tissue‐visualization advantages of each individual mode (e.g. similar to polarized and non‐polarized dermoscopy complementing each other) . The importance of the novel FuCM device is that it improves the visualization of nuclear, cytoplasmic and stromal details by simultaneously obtaining the three CM modes: RCM, FCM and FuCM.…”