Tomography is a powerful tool for obtaining three-dirotation geometry. Although single-axis data collection is stanmensional information from transmission electron microscopy, but its dard in biomedical imaging (e.g., CAT scan, scanning via compuapplication faces unique challenges. A single-axis tilt geometry for terized axial tomography), other data collection geometries are data collection results in anisotropic resolution because full angular also routinely used. coverage is not feasible for most specimen preparations. This effect In the last 10 years, electron tomography has proved to be an can be minimized by combining two single-axis tilt series that have effective tool for 3D structure–function determination in cellular been collected orthogonal to each other. Rapid freezing has been and molecular biology (reviewed in [6–8]) . Recent successful successfully used to preserve the native structure of biological speciapplications include determining the location of microtubule initimens in a form that can be visualized in the high-vacuum environment ation sites in centrosomes isolated from fruit fly embryos [9] , required for electron microscopy; however, these preparations are extremely labile to electron exposure. As a result, application of todemonstrating dynamic changes that occur in muscle Z bands mography to frozen-hydrated specimens has only recently become with contraction [10], characterizing the arrangement and morfeasible with the development of automatic data collection, and with phology of crystals formed during the initial events of bone fora renewed appreciation for the principle of dose fractionation. Even mation [11,12], and providing new insight into the folding of with the limitations of traditional specimen preparations and convenDNA into chromosomes [13,14]. Here we briefly review the tional methods of data collection, electron microscopic tomography methodology, including recent efforts to overcome radiation damhas been successfully employed to probe the structure and function age and an unfavorable specimen geometry, and present a new of several important cellular components. Current efforts include comapplication where electron tomography is combined with videobining electron microscopic tomography and video-enhanced light enhanced light microscopy to correlate structural determination microscopy to correlate ultrastructural variation with the direction of with motion. chromosome motion during mitosis. q 1997 John Wiley & Sons, Inc. Int J Imaging Syst Technol, 8, 175–187, 1997