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Skeletal metastasis is common in patients with advanced breast cancer and often caused by immune evasion of disseminated tumor cells (DTCs). In the skeleton, tumor cells not only disseminate to the bone marrow but also to osteogenic niches in which they interact with newly mineralizing bone extracellular matrix (ECM). However, it remains unclear how mineralization of collagen type I, the primary component of bone ECM, regulates tumor‐immune cell interactions. Here, a combination of synthetic bone matrix models with controlled mineral content, nanoscale optical imaging, and flow cytometry are utilized to evaluate how collagen type I mineralization affects the biochemical and biophysical properties of the tumor cell glycocalyx, a dense layer of glycosylated proteins and lipids decorating their cell surface. These results suggest that collagen mineralization upregulates mucin‐typeO‐glycosylation and sialylation by tumor cells, which increases their glycocalyx thickness while enhancing resistance to attack by natural killer (NK) cells. These changes are functionally linked as treatment with a sialylation inhibitor decreased mineralization‐dependent glycocalyx thickness and made tumor cells more susceptible to NK cell attack. Together, these results suggest that interference with glycocalyx sialylation may represent a therapeutic strategy to enhance cancer immunotherapies targeting bone‐metastatic breast cancer.

Azimuthal beam scanning eliminates the uneven excitation field arising from laser interference in through-objective total internal reflection fluorescence (TIRF) microscopy. The same principle can be applied to scanning angle interference microscopy (SAIM), where precision control of the scanned laser beam presents unique technical challenges for the builders of custom azimuthal scanning microscopes. Accurate synchronization between the instrument computer, beam scanning system and excitation source is required to collect high quality data and minimize sample damage in SAIM acquisitions. Drawing inspiration from open-source prototyping systems, like the Arduino microcontroller boards, we developed a new instrument control platform to be affordable, easily programmed, and broadly useful, but with integrated, precision analog circuitry and optimized firmware routines tailored to advanced microscopy. We show how the integration of waveform generation, multiplexed analog outputs, and native hardware triggers into a single central hub provides a versatile platform for performing fast circle-scanning acquisitions, including azimuthal scanning SAIM and multiangle TIRF. We also demonstrate how the low communication latency of our hardware platform can reduce image intensity and reconstruction artifacts arising from synchronization errors produced by software control. Our complete platform, including hardware design, firmware, API, and software, is available online for community-based development and collaboration.