Confocal Examination of Nonmelanoma Cancers in Thick Skin Excisions to Potentially Guide Mohs Micrographic Surgery Without Frozen Histopathology

Abstract

Precise removal of nonmelanoma cancers with minimum damage to the surrounding normal skin is guided by the histopathologic examination of each excision during Mohs micrographic surgery. The preparation of frozen histopathology sections typically requires 20-45 min per excision. Real-time confocal reflectance microscopy offers an imaging method potentially to avoid frozen histopathology and prepare noninvasive (optical) sections within 5 min. Skin excisions ( approximately 1 mm thick) from Mohs surgeries were washed with 5% acetic acid and imaged with a confocal cross-polarized microscope. The confocal images were compared with the corresponding histopathology. Acetic acid causes compaction of chromatin that increases light back-scatter and makes the nuclei bright and easily detectable. Crossed-polarization strongly enhances the contrast of the nuclei because the compacted chromatin depolarizes the illumination light whereas the surrounding cytoplasm and normal dermis does not. Fast low-resolution examination of cancer lobules in wide fields of view followed by high-resolution inspection of nuclear morphology in small fields of view is possible; this is similar to the procedure for examining histopathology sections. Both the Mohs surgeon and the patient will potentially save several hours per day in the operating room. Fast confocal reflectance microscopic examination of excisions (of any thickness) may improve the management of surgical pathology and guide microsurgery of any human tissue.

Review paper The use of reflectance confocal microscopy in selected inflammatory skin diseases

“…It has a wide range of applications, particularly in the diagnosis of neoplastic skin lesions, enabling non-invasive monitoring of treatment [2][3][4][5][6], in examining skin reactions to external factors (e.g. ultraviolet radiation) [7], as well as planning surgical margins in the course of pre-and intraoperative evaluation [8][9][10]. Reflectance confocal microscopy has also been applied in the study of a number of non-neoplastic skin diseases, especially inflammatory dermatoses.…”

Modulated-alignment dual-axis (MAD) confocal microscopy for deep optical sectioning in tissues

“…The ability to visualize these tissues clearly and to investigate these processes of cancer invasion would benefit from a technology capable of imaging intact tissues to a depth of 500 μm or more. The need for high spatial resolution (super-resolution microscopy [5][6][7][8]) in clinical diagnoses is less immediate since the "gold standard" of histopathology relies on the observation of tissue sections under low magnification (e.g., 2x -10x is typical for Mohs surgical pathology) [9]. Rather, for applications such as early cancer detection, guided biopsy (e.g., in oral cancer), or surgical guidance (e.g., in dermatology), the ability to visualize large fields of view in three dimensions with cellular resolution is valuable, especially if there is sufficient depth to enable assessment of the local invasion of cancer.…”

Contrast improvement in scattered light confocal imaging of skin birefringent structures by depolarization detection

“…Confocal laser scanning microscopy is a reliable and robust technique to provide non-invasive realtime optical imaging of biological tissue at high resolution and contrast, approaching histology, the golden standard for tissue inspection [12][13]. The detection of cross-polarized component of light can be used to enhance the intensity of light reflected or scattered from the deeper tissue layers, while rejecting direct reflectance from its surface [13].…”

“…The detection of cross-polarized component of light can be used to enhance the intensity of light reflected or scattered from the deeper tissue layers, while rejecting direct reflectance from its surface [13].…”

“…In the past decade the interest in studying the propagation of polarized light in birefringent media has significantly increased [8][9]. Polarization-based theoretical methods, such as timeresolved [10] and Mie theory-based [11] Monte Carlo algorithms, and experimental techniques, such as polarization-sensitive optical coherence tomography (PS-OCT) [1][2][3][4] and cross-polarized confocal laser scanning microscopy (CPCLSM) [12][13] have advanced the understanding of birefringence in biological tissues. These theoretical and experimental tools were used to characterize a change in polarization of light due to birefringent properties of biological tissues.…”