During immune surveillance, cytotoxic T lymphocytes (CTL) can selectively identify and destroy tumor cells by recognizing tumor-specific peptides (neoantigens), bound to major histocompatibility complex molecules (pMHC) arrayed on cancer cell surfaces. CTL use the same machinery to destroy virally infected cells displaying pathogen-specific pMHC, while leaving intact healthy cells expressing normal self-pMHC. We present a robotic microscope that allows scientists to conduct highly sensitive and

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... selective T cell-pMHC studies with high throughput. Our system manipulates micro-meter beads coated with particular pMHC, presents them to T cells and generates piconewtonlevel intermolecular forces required to detect T cell acuity with a neoantigen. Our system integrates optical tweezers, precision nanomicro stages, and episcopic/diascopic illumination schemes at two magnifications. We create a coordinate referencing system to locate translucent T cells in three-dimensional, over a large space, based on the characteristic intensity change at each focal plane caused by the cells. Our system performs automated experiments to detect the level of T cell acuity with specific pMHCs, by measuring the downstream cellular responses. High acuity T cells can be selectively recovered for single cell analysis. Our new methodology and tool will have a significant impact on cancer immunotherapy and immunology research. Index Terms-Automation at micro-nano scales, automation in life sciences, biotechnology, pharmaceutical and health care, biological cell manipulation.