High-content stimulated Raman histology of human breast cancer.

Histological examination is crucial for cancer diagnosis, however, the labor-intensive sample preparation involved in the histology impedes the speed of diagnosis. Recently developed two-color stimulated Raman histology could bypass the complex tissue processing to generates result close to hematoxylin and eosin staining, which is one of the golden standards in cancer histology. Yet, the underlying chemical features are not revealed in two-color stimulated Raman histology, compromising the effectiveness of prognostic stratification. Here, we present a high-content stimulated Raman histology (HC-SRH) platform that provides both morphological and chemical information for cancer diagnosis based on un-stained breast tissues. Methods: By utilizing both hyperspectral SRS imaging in the C-H vibration window and sparsity-penalized unmixing of overlapped spectral profiles, HC-SRH enabled high-content chemical mapping of saturated lipids, unsaturated lipids, cellular protein, extracellular matrix (ECM), and water. Spectral selective sampling was further implemented to boost the speed of HC-SRH. To show the potential for clinical use, HC-SRH using a compact fiber laser-based stimulated Raman microscope was demonstrated. Harnessing the wide and rapid tuning capability of the fiber laser, both C-H and fingerprint vibration windows were accessed. Results: HC-SRH successfully mapped unsaturated lipids, cellular protein, extracellular matrix, saturated lipid, and water in breast tissue. With these five chemical maps, HC-SRH provided distinct contrast for tissue components including duct, stroma, fat cell, necrosis, and vessel. With selective spectral sampling, the speed of HC-SRH was improved by one order of magnitude. The fiber-laser-based HC-SRH produced the same image quality in the C-H window as the state-of-the-art solid laser. In the fingerprint window, nucleic acid and solid-state ester contrast was demonstrated. Conclusions: HC-SRH provides both morphological and chemical information of tissue in a label-free manner. The chemical information detected is beyond the reach of traditional hematoxylin and eosin staining and heralds the potential of HC-SRH for biomarker discovery.

Millimeter-deep micron-resolution vibrational imaging by shortwave infrared photothermal microscopy.

Deep-tissue chemical imaging plays a vital role in biological and medical applications. Here, we present a shortwave infrared photothermal (SWIP) microscope for millimeter-deep vibrational imaging with sub-micron lateral resolution and nanoparticle detection sensitivity. By pumping the overtone transition of carbon-hydrogen bonds and probing the subsequent photothermal lens with shortwave infrared light, SWIP can obtain chemical contrast from polymer particles located millimeter-deep in a highly scattering phantom. By fast digitization of the optically probed signal, the amplitude of the photothermal signal is shown to be 63 times larger than that of the photoacoustic signal, thus enabling highly sensitive detection of nanoscale objects. SWIP can resolve the intracellular lipids across an intact tumor spheroid and the layered structure in millimeter-thick liver, skin, brain, and breast tissues. Together, SWIP microscopy fills a gap in vibrational imaging with sub-cellular resolution and millimeter-level penetration, which heralds broad potential for life science and clinical applications.