Characteristics of coherent gating technique are inherently well-suited for ideal histopathologic imaging, and it is particularity useful as an alternative to histological tissue observation when it is not practical to take specimens for histological processing or when large areas of tissue are needed to be investigated. In particular, tomographic imaging has an important role as optical biopsy, because it can provide a real-time structural information non-invasively at spatial resolutions similar to that of a standard histological section taken from a biopsy sample. To date, optical imaging based on coherent gating technique has widely been applied in the clinic to diagnose diseases of various tissues 27, 28. OCM is also suitable for deep tissue imaging, because OCM uses near infrared light that reaches deeper into biological tissues than visible light can reach. Thus, it does not require chemical labeling, staining or external contrast agents. OCM is based on a coherent gating technique of optical interferometry, and utilizes the endogenous back-scattering signal. Optical coherence microscopy (OCM) 22, 23, 24, 25, 26 could be an alternative imaging modality for high throughput histopathology. An ideal 3D optical imaging technique for histopathology would be able to image unlabeled and unstained fresh specimens and provide high throughput quantitative information as well as native 3D context. However, it requires careful sample preparation for clearing protocols as well as long incubation for immunostaining, which can often distort the physical volume of the entire 3D specimens and thus can restrict its utility for pathology. This approach allows fast volumetric imaging without the need for mechanical sectioning. A number of tissue clearing techniques 14, 19 have been introduced and successfully combined with light-sheet microscopy to unveil the cellular connectivity at large areas. As the effective clearing agents are available, the deep tissue optical imaging has become feasible through either the refractive index matching of tissue or removal of the highly scattered biological components like lipid. While these techniques can generate a complete dataset such as projectomes and connectomes, the high cost of equipment, the huge amount of time needed for tissue processing and fluorescent labeling, and massive data collection and signal processing make this technique difficult to be widely available for pathology applications.Īnother novel approach for volumetric imaging is based on light-sheet fluorescence microscopy and chemically clearing tissue 14, 15, 16, 17, 18, 19, 20, 21. One of representative attempts of fully automated bright field microscopy 11, single photon microscopy 12 and two-photon microscopy combined with tissue sectioning method 13 were successfully used to reconstruct fine neuronal network and anatomical tracings in a whole mouse brain. 3D microscopic imaging techniques have been initially applied to neuroscience and developmental studies. Over the past decade, advanced 3D microscopic imaging techniques for high resolution and volumetric anatomy have been demonstrated. However, current methods lack the ability to visualize diagnostic pathologic feature over large areas or volumetric context. In order to overcome current restriction of histopathology, several optical imaging techniques have been introduced toward rapid and stain-free histopathology of fresh tissues 4, 5, 6, 7, 8, 9, 10. Although histologic analysis in pathology is a gold-standard to understand tissue morphology, diagnose diseases, and decide treatment course, it is still very time- and labor-intensive and provides only limited tissue information in thin and narrow field of view. The histological examination of excised tissue via optical microscope has been widely used in biological laboratory and clinic, which requires the cumbersome process such as fixation, embedding, sectioning, and staining of biological specimens 1, 2, 3.
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