Fast and Accurate Fiducial Marker Tracking with Fluorescence Microscopy for Mechanobiology Experiments

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Equipment is being requested to support research activities at Carleton University focused on understating how mechanical strain is sustained by and transmitted through biological tissues. The equipment will enable rapid three-dimensional (3D) tracking of fluorescent fiducial markers embedded on soft tissue specimens in response to mechanical stimulation with unprecedented levels of both spatial (~20nm) and temporal resolution (~2ms). Insights gained from this experimental system will lead to a better understanding of the key structures that determine tissue mechanical properties and the mechanisms of tissue injury in mild traumatic brain injury and dermopathies. The speed and resolution of the equipment is achieved through a point spread function engineering module (Double Helix phase mask) and a high-speed camera (Photometrics Kinetix). This system allows 3D displacements of fluorescent fiducial markers to be measured without physically adjusting the focal plane of the microscope, and therefore measurements can be made much faster than previously attainable (~seconds). Using this new technology to measure the deformation of sliced tissue in response to mechanical stimulation is a new concept and holds the potential for rapid advancement of the cell and tissue mechanics fields, where high strain rate measurements have been challenging. In addition, this system offers the opportunity to perform 3D super-resolution imaging as an enhancement to a recently funded Total Internal Reflection Fluorescence microscope that will be installed at Carleton University. Point spread function engineering of this kind is rapidly being adopted for super-resolution imaging, but there is currently limited equipment available in Canada. The requested equipment will directly support broad research efforts pertaining to tissue mechanics at Carleton University including the Tissue Engineering and Applied Materials (TEAM hub) and the Ottawa Carleton Institute for Biomedical Engineering (OCIBME) and therefore will be of immediate benefit to 5 research groups and their HQP. The applicants are experts in the synergistic fields of tissue engineering, material mechanics and optical microscopy, forming an ideal team will make full use of the requested equipment. The lead applicant has extensive experience in single molecule imaging, cell, and tissue mechanics and therefore the equipment will be used immediately and effectively. To continue to compete internationally and provide proper training to HQP, sustained access to interdisciplinary tools such as cutting-edge optical microscopy and sample preparation techniques are essential, without which it will be hard to maintain a respectable standing within our research community. Three current graduate students and two undergraduate researchers stand to benefit immediately from the use of this equipment, a number that is anticipated to grow as the requested equipment enables new lines of inquiry to develop.