Three-dimensional reconstruction of histological serial sections on the computer

This paper reports the development of an objective and time-saving method for the three-dimensional reconstruction of sectioned morphological specimens on the computer. The development of a fiducial marker technique, the design of program extensions for the automatic thresholding and realignment of images, and a technique for the transfer of marker-derived reference points into images of higher magnification are described. The method is based on the use of resin as embedding material and drill holes as fiducials. Images are digitized by a video camera, and NIH-Image is used for image processing. Special macros were written for the automation of a number of operations, such as object and marker segmentation, image reorientation, image scaling, dirt removal, and marker transfer. The so-treated image series were transferred to a workstation and were reconstructed using ''Image Volumes''. The reconstructs obtained from this largely automated technique exhibit a perfect alignment of the sections. The 3-D representations of the sectioned objects have a much higher degree of objectivity than those obtained by manual image orientation or by semiautomated techniques that do not use fiducials.