Fast wavelength switching controls and a high sensitivity camera for enhanced imaging

Whether a cell is free-living in the environment or part of a layer of cells surrounding an organ or tissue, it must interact and sense its surroundings. Each cell is contained within a bilayer of fats (or lipids) and proteins referred to as the plasma membrane. Interactions between a cell and its surroundings or neighboring cells occur through the proteins and lipids located on the cell's outer surface. The plasma membrane and its many different surface proteins allow each cell to take up or "eat" nutrients such as sugars, dietary fat, and vitamins. The plasma membrane also allows a cell to communicate with other cells by receiving signals in the form of hormones and forming protective barriers such as skin or kidneys. Many of these features are examined and studied by the applicants under the umbrella of cell biology. As cell biologists, it is of paramount importance for us to visualize not only the processes under examination but also the molecules that control these events. In our research, that means visualizing the membrane proteins and lipids that mediate cellular organization, movement, and housekeeping.

Epifluorescence microscopy is a fluorescence imaging technique that relies on illumination of the entire field of view of a fluorescence microscope and simultaneous detection of the emitted fluorescence using a camera. Due to the nature of this system, the camera collects thick sections of the specimen including light that is out of focus. However, this technique is useful when monitoring diffuse signals within the cell such as probes to monitor calcium or acidity. To enable optimal performance of this equipment fast shutters and switching of wavelengths are required. The current epifluorescent system contains only manual shutters and wavelength switching that severely hampers its usefulness and ability to conduct experiments. The first item we request is the necessary components to upgrade our epifluorescence microscope to greatly increase productivity and throughput.

To complement epifluorescence microscopes, researchers have developed the confocal microscope. In these systems much of the emitted light is filtered resulting in the ability to examine finer structures and collect thinner samples. One trade off with the filtering unwanted light is loss of signal. Thus, the second upgrade we are requesting is a camera for a spinning disc confocal with enhanced sensitivity due to a vastly superior efficiency of light capture.

As such, NSERC funding is essential to purchase upgrades for two existing microscopes to enhance greatly their usability and usefulness that will be critical for many research programs and trainees at St. Michael’s Hospital.