title: Confocal DNA cytometry: a contour-based segmentation algorithm for automated three-dimensional image segmentation.
abstract: BACKGROUND
  Confocal laser scanning microscopy (CLSM) presents the opportunity to perform three-dimensional (3D) DNA content measurements on intact cells in thick histological sections. So far, these measurements have been performed manually, which is quite time-consuming.
  
  
  METHODS
  In this study, an intuitive contour-based segmentation algorithm for automatic 3D CLSM image cytometry of nuclei in thick histological sections is presented. To evaluate the segmentation algorithm, we measured the DNA content and volume of human liver and breast cancer nuclei in 3D CLSM images.
  
  
  RESULTS
  A high percentage of nuclei could be segmented fully automatically (e.g., human liver, 92%). Comparison with (time-consuming) interactive measurements on the same CLSM images showed that the results were well correlated (liver, r = 1.00; breast, r = 0.92).
  
  
  CONCLUSIONS
  Automatic 3D CLSM image cytometry enables measurement of volume and DNA content of large numbers of nuclei in thick histological sections within an acceptable time. This makes large-scale studies feasible, whereby the advantages of CLSM can be exploited fully. The intuitive modular segmentation algorithm presented in this study detects and separates overlapping objects, also in two-dimensional (2D) space. Therefore, this algorithm may also be suitable for other applications.
authors.affiliation: Department of Pathology, VU Medical Center, Amsterdam, The Netherlands. jam.belien@vumc.nl
authors.forename: Jeroen A M
authors.initials: JA
authors.lastname: Beliën
authors.forename: Hielke A H M
authors.initials: HA
authors.lastname: van Ginkel
authors.forename: Paulos
authors.initials: P
authors.lastname: Tekola
authors.forename: Lennert S
authors.initials: LS
authors.lastname: Ploeger
authors.forename: Neal M
authors.initials: NM
authors.lastname: Poulin
authors.forename: Jan P A
authors.initials: JP
authors.lastname: Baak
authors.forename: Paul J
authors.initials: PJ
authors.lastname: van Diest
mesh_terms: D000465:Algorithms; D001943:Breast Neoplasms / Q000473:pathology*; D002467:Cell Nucleus / Q000473:pathology; D004273:DNA, Neoplasm / Q000032:analysis; D005260:Female; D006801:Humans; D019044:Image Cytometry / Q000295:instrumentation / Q000379:methods*; D007091:Image Processing, Computer-Assisted / Q000295:instrumentation / Q000379:methods*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008113:Liver Neoplasms / Q000473:pathology*; D018613:Microscopy, Confocal / Q000379:methods*; D011003:Ploidies
chemical_list: D004273:DNA, Neoplasm