A BioAFM for Intracellular, Cellular, and Tissue Measurement and Manipulation

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To deepen our understanding of the mechanics and functions of cells, intracellular organelles and tissues, new techniques are required, including direct mechanical measurement and manipulation.******This RTI application requests a biological atomic force microscope (bioAFM) to support this team of researchers to explore the mechanics and mechanobiology of cells, intracellular structures, and tissues. Different from previous models of bioAFMs, this requested bioAFM contains the following unique capabilities, which are essential to our proposed research programs. (1) A FluidFM module for the extraction of picolitre and nanoliter subcellular substances (e.g., biopsy of mitochondria, mRNA, nnucleic acids etc.). (2) Fast scanning and indention that enable capturing dynamics of cellular activities, such as the sealing of the cell membrane, the protein structure change on the nuclear pore complex before and after mechanical indentation, and the activation of mechanosensitive ion channels (e.g., PIEZO1). (3) A large Z range that is critical for scanning rough surfaces such as vibratome-sectioned tissue slices. (4) An open-stage incubation chamber that controls and maintains the cell incubation environment (e.g., temperature and CO2) while permitting manipulation tools (e.g., AFM probe and glass micropipette) for sample access. ******The requested bioAFM will be mounted on our existing confocal microscope (for 3D high-resolution fluorescence imaging) and integrated with our precision cell ablation laser (for opening a hole on cell membrane or ablate a filament in a cell) to form a powerful system. Our HQP will use the equipment to conduct research on intracellular, cellular and intra-tissue mechanics and mechanobiology experiments and make exciting discoveries. The equipment will be used to train a high number of postdocs and graduate students who will gain skills in cell and tissue mechanics, cell mechanotransduction, cell manipulation, development biology, microscopy imaging, and precision instrument control and measurement.