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Cellular biomechanics plays critical roles in cancer metastasis and tumor progression. Existing studies on cancer cell biomechanics are mostly conducted in flat 2D conditions, where cells' behavior can differ considerably from those in 3D physiological environments. Despite great advances in developing 3D in vitro models, probing cellular elasticity in 3D conditions remains a major challenge for existing technologies. In this work, we utilize optical Brillouin microscopy to longitudinally acquire mechanical images of growing cancerous spheroids over the period of eight days. The dense mechanical mapping from Brillouin microscopy enables us to extract spatially resolved and temporally evolving mechanical features that were previously inaccessible. Using an established machine learning algorithm, we demonstrate that incorporating these extracted mechanical features significantly improves the classification accuracy of cancer cells, from 74% to 95%. Building on this finding, we have developed a deep learning pipeline capable of accurately differentiating cancerous spheroids from normal ones solely using Brillouin images, suggesting the mechanical features of cancer cells could potentially serve as a new biomarker in cancer classification and detection., Competing Interests: Conflict of Interest The authors declare no conflict of interest.

Brillouin microscopy based on spontaneous Brillouin scattering has emerged as a unique elastography technique because of its merit of non-contact, label-free, and high-resolution mechanical imaging of biological cell and tissue. Recently, several new optical modalities based on stimulated Brillouin scattering have been developed for biomechanical research. As the scattering efficiency of the stimulated process is much higher than its counterpart in the spontaneous process, stimulated Brillouin-based methods have the potential to significantly improve the speed and spectral resolution of existing Brillouin microscopy. Here, we review the ongoing technological advancements of three methods, including continuous wave stimulated Brillouin microscopy, impulsive stimulated Brillouin microscopy, and laser-induced picosecond ultrasonics. We describe the physical principle, the representative instrumentation, and biological application of each method. We further discuss the current limitations as well as the challenges for translating these methods into a visible biomedical instrument for biophysics and mechanobiology., Competing Interests: Conflict of interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.