MEASUREMENT BY CONFOCAL LASER-SCANNING MICROSCOPY OF THE VOLUME OF EPIDERMAL NUCLEI IN THICK SKIN SECTIONS

The mean volume and shape of nuclei assessed in standard tissue sectio ns by means of stereologic and morphometric methods are associated wit h prognosis in tumors of different sites. Thus, accurate quantificatio n of the volume and shape of cell nuclei can be important for cancer p atients and also may be useful for a better understanding of basic cel lular events, such Its growth and differentiation. Confocal laser scan microscopy (CLSM) makes it possible to obtain image sets consisting o f very thin serial optical slices from thick tissue sections. These im ages can be used with digital image processing to construct a three-di mensional (3-D) model of individual nuclei. We used CLSM to study a 50 -mu m-thick paraffin section of a skin biopsy. Image processing was ap plied to the CLSM images to precisely segment epidermal nuclei from th e background, and serial two-dimensional (2-D) binary images were crea ted. Alignment of the 2-D images that form the 3-D model of the origin al nuclei was carefully controlled. The series of 2-D binary images wa s connected with an algorithm to form the 3-D model of each complete n ucleus and organized for the 3-D visualization and analysis using a 3- D volume rendering method. In this way we measured the volume and surf ace area of 22 intact epidermal nuclei. The mean nuclear volume was 17 4.7 mu m(3), the standard deviation of the volume 26.47 mu m(3), the m ean surface area 168.0 mu m(2) and the standard deviation of the surfa ce area 22.00 mu m(2). At this time the procedure is time consuming du e to the prototype research nature of the image processing program. Ho wever, it is expected that with fairly simple improvements in the soft ware, the speed can increase to make the technique of practical value in surgical pathology.