Knife-edge scanning microscopy for imaging and reconstruction of three-dimensional anatomical structures of the mouse brain.
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abstract
Anatomical information at the cellular level is important in many fields, including organ systems development, computational biology and informatics. Creating data sets at resolutions that provide enough detail to reconstruct cellular structures across tissue volumes from 1 to 100 mm(3) has proven to be difficult and time-consuming. In this paper, we describe a new method for staining and imaging large volumes of tissue at sub-micron resolutions. Serial sections are cut using an automated ultra-microtome, whereas concurrently each section is imaged through a light microscope with a high-speed line-scan camera. This technique, knife-edge scanning microscopy, allows us to view and record large volumes of tissue in a relatively small amount of time (approximately 7 mm(2) s(-1)). The resolution and scanning speed of knife-edge scanning microscopy provides a new method for imaging tissue at sufficient resolution to reconstruct maps of cellular distribution and morphology. We show that these techniques preserve the alignment of serial sections accurately enough to allow for reconstruction of neuronal processes and microvasculature. Expanding these techniques to other tissues opens up the possibility of creating fully reconstructed cellular maps of entire organs.
