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This review examines the clinical application of label-free microscopy and spectroscopy, which are based on optical signals emitted by tissue components. Over the past three decades, a variety of techniques have been investigated with the aim of developing an in situ histopathology method that can rapidly and accurately identify tumor margins during surgical procedures. These techniques can be divided into two groups. One group encompasses techniques exploiting linear optical signals, and includes infrared and Raman microspectroscopy, and autofluorescence microscopy. The second group includes techniques based on nonlinear optical signals, including harmonic generation, coherent Raman scattering, and multiphoton autofluorescence microscopy. Some of these methods provide comparable information, while others are complementary. However, all of them have distinct advantages and disadvantages due to their inherent nature. The first part of the review provides an explanation of the underlying physics of the excitation mechanisms and a description of the instrumentation. It also covers endomicroscopy and data analysis, which are important for understanding the current limitations in implementing label-free techniques in clinical settings. The second part of the review describes the application of label-free microscopy imaging to improve oncological surgery with focus on brain tumors and selected gastrointestinal cancers, and provides a critical assessment of the current state of translation of these methods into clinical practice. Finally, the potential of confocal laser endomicroscopy for the acquisition of autofluorescence is discussed in the context of immediate clinical applications.
Galli et al. (Thu,) studied this question.