Optimization of a Two-Photon Laser Scanning Microscope, Poster 17

Imagine being able to look inside a sample layer by layer and observe details on the cellular level, without making an incision. Optical sectioning allows images to be taken at different depths, and then stacked to create a... [ view full abstract ]

Imagine being able to look inside a sample layer by layer and observe details on the cellular level, without making an incision. Optical sectioning allows images to be taken at different depths, and then stacked to create a three-dimensional representation of the sample. Our laser-scanning microscope takes advantage of two-photon excitation to obtain optical sectioning. By focusing the beam so that only a small point fluoresces and then scanning this point around the sample, our microscope constructs three-dimensional images. I rebuilt components of the laser-scanning microscope, incorporating a shutter to block the beam between exposures, improving motorized control of the sample stage, and mitigating power loss between the laser source and the sample. I also realigned our wide-field camera so that it can be used as a reference, ensuring that the sample is placed at the focus of the laser. Because of these modifications, the microscope can easily image biological samples for the first time. We will present images of Drosophila melanogaster (fruit fly) brain cells expressing autofluorescence. Current work includes optimizing laser excitation and emission detection wavelengths for biological samples and increasing sample rate to improve image quality.