ABSTRACT While our ability to detect and treat primary tumors has significantly increased in the past decade, it remains difficult to diagnose the origination of metastasis which remains responsible for nearly 90% of cancer-related deaths. In this respect, recently, the critical role of the mechanical state of tumors and tumor cells has been recognized for tumor progression, malignancy transformation and metastasis but it remains poorly exploited due to the lack of suitable measurement tools. Many microfluidic deformability approaches have recently emerged, but they only give an average value of the overall cell mechanical properties, while it would be important to separate nucleus vs cytoskeleton contributions, and they need mechanical stimulation to probe properties, which is deleterious since cells strongly react to mechanical stimuli. In the past few years, we have been developing an all-optical approach to this challenge, named Brillouin microscopy, and strongly established it in tumor biology to characterize cell mechanics during metastatic cascade. However, current technology is inherently limited in speed (~50ms/point), as it relies on spontaneous Brillouin interaction, and thus is not suitable for rapid screening/sorting of tumor cells. Here, we will develop stimulated Brillouin cytometry which 1) increases speed by ~100-fold and 2) provides additional contrast mechanisms such as viscosity and mass density with micron-scale resolution. Based on this breakthrough, we will develop and validate a flow cytometry/cell sorting platform using elastic modulus, viscosity, and density as label-free contrast mechanisms (Aim 1). We will then validate our mechanical assessment of metastatic cells against microfluidic assays to assess migration and proliferation and in mice models to assess cell’s metastatic ability in vivo (Aim 2). The rigorous process of technology development, validation, benchmarking and field-testing will yield a platform with unprecedented capabilities to characterize and sort cells based on their mechanical properties within an instrument compatible with traditional flow cytometry, thus ready to be widely adopted by the cancer biology community. The proposal features the collaboration between optical technology experts (UMD) and cancer metastasis pioneers in both in vitro (JHU) and in vivo (UMB) settings with an established track record of fruitful collaboration.