Multi-laser Total Internal Reflection Fluorescence Microscopy

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Total Internal Reflection Fluorescence (TIRF) Microscopy is a recent innovation in the field of fluoresence imaging. Researchers can now examine the behaviour of single particles, such as the movement of motor proteins on filamentous tracks or to measure the activity of cellular components in living cells by tagging their objects of interest with fluorescent proteins. With TIRF, only a very thin layer of the sample is exposed to the light that causes fluorescent output, resulting in a better ability to distinguish individual fluorescent objects, and the capacity to image specimens with low excitation over long periods of time. This proposal will bring a recently purchased basic TIRF system to full operating capacity through the acquisition of 3 additional lasers and a camera that will enable simultaneous imaging of multiple fluorescent objects. Researchers with a wide range of applications and backgound will benefit from this new facility. CRC Geoffrey Wasteneys is uncovering the mechanisms that drive the growth and shrinkage of dynamic filaments known as microtubules, which form form micro-machines critical for cellular processes. His team will use the TIRF system to observe the behaviour of proteins that associate with and control the growth and shrinkage of these microtubule polymers. Mathematician Eric Cytrynbaum will use TIRF data as inputs for computational simulations, to model in mathematical terms the processes of self organization that generate patterns of subcellular organization. Lacey Samuels' laboratory will use the TIRF system to study how lipids are secreted and recycled in plant cells, a little understood process critical for plant growth and development. For Jay Kizhakkedathu's research team, TIRF is the only known method by which they can monitor how cells adhere to and proliferate with the bioactive surfaces they are developing, a process that will optimizing drug delivery and implants. Nancy Forde, a biophysicist at SFU is part of an international team attempting to produce the first autonomous, protein-based synthetic molecular motor. For them, TIRF is critical for visualising and quantifying the activity of their prototype motors.