Multiple Imaging Modes for Enhanced Analysis of Drosophila Cells with the Leica SP8 Confocal System

Multiple Imaging Modes for Enhanced Analysis of Drosophila Cells and Tissues with the Leica SP8 Confocal Microscope System (2022 Poster Presentation, Kevin Edwards). The Confocal Microscopy Facility at Illinois State University is open to regional users and features several capabilities that allow enhanced characterization of fly tissues. 1) Resolution enhancement. We extensively tested Leica's Lightning adaptive deconvolution tool and found it greatly improved imaging of defined objects on the 0.1-1 micron scale such as protein aggregates or cytoskeletal structures. 2) Spectral analysis. Certain challenging problems such as colocalization of overlapping dyes, and precisely defining shifts in dye fluorescent properties, can be solved using "spectral fingerprinting" (Lambda-lambda plots) in which every excitation/emission combination is imaged and graphically analyzed. 3) Fluorescence lifetime imaging (FLIM). The timing of fluorescent emissions can be used to complement spectral analysis, providing a way to distinguish different chemical species with similar spectra. We find, for example, that FLIM analysis of cuticle autofluorescence can distinguish among functionally different antennal sensilla.