DNA QUANTIFICATION IN CERVICAL INTRAEPITHELIAL NEOPLASIA THICK TISSUE-SECTIONS BY CONFOCAL LASER-SCANNING MICROSCOPY

Image analysis of tissue biopsies for determination of DNA content as an early marker of neoplasia is hampered by the complexity of correcti ons necessary to deal with nuclear truncation and overlap in thin sect ions. The use of confocal laser scanning microscopy (CLSM) for measure ment of cellular DNA content on whole cells within thick tissue sectio ns offers the advantage of preservation of cellular architecture, capa city for 3-dimensional analysis, and absence of sectioning artifacts. We have applied this technique to pararosaniline-feulgen stained human cervical tissues graded from normal to cervical intraepithelial neopl asia (GIN) III. For the purpose of comparison, 15 mu m sections were s tained and mapped so that the same cell population could be analyzed b y both integrated optical density and fluorescence intensity. Distribu tion of DNA content from normal cervical epithelial cells 2-3 layers o ut from the basal cell layer measured by both methodologies showed a s table G0/G1 population with no observable S-phase or G2 cells. Cells m easured from areas of increasing CIN grade showed progressively higher DNA content values that were not observable in normal tissue. Althoug h these data are preliminary they suggest that CLSM can be used to ide ntify aneuploid states within defined structural areas of pre-invasive neoplasia. (C) 1997 Wiley-Liss, Inc.