Semi-Automated Reconstruction of Neural Processes from Large Numbers of Fluorescence Images
Fiala, John C.
Lichtman, Jeff W.
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Citation (published version)Lu, Ju, John C. Fiala, Jeff W. Lichtman. "Semi-Automated Reconstruction of Neural Processes from Large Numbers of Fluorescence Images" PLoS ONE 4(5): e5655. (2009)
We introduce a method for large scale reconstruction of complex bundles of neural processes from fluorescent image stacks. We imaged yellow fluorescent protein labeled axons that innervated a whole muscle, as well as dendrites in cerebral cortex, in transgenic mice, at the diffraction limit with a confocal microscope. Each image stack was digitally re-sampled along an orientation such that the majority of axons appeared in cross-section. A region growing algorithm was implemented in the open-source Reconstruct software and applied to the semi-automatic tracing of individual axons in three dimensions. The progression of region growing is constrained by user-specified criteria based on pixel values and object sizes, and the user has full control over the segmentation process. A full montage of reconstructed axons was assembled from the ~200 individually reconstructed stacks. Average reconstruction speed is ~0.5 mm per hour. We found an error rate in the automatic tracing mode of ~1 error per 250 um of axonal length. We demonstrated the capacity of the program by reconstructing the connectome of motor axons in a small mouse muscle.