Automated topographical cell proliferation analysis.
BACKGROUND
Cell proliferation is often studied using the incorporation of bromodeoxyuridine (BrdU). Immunohistochemical staining is then used to detect BrdU in the nucleus. To circumvent the observer bias and labor-intensive nature of manually counting BrdU-labeled nuclei, an automated topographical cell proliferation analysis method is developed.


METHODS
Sections stained with fluorescein-labeled anti-BrdU and counterstained with To-Pro-3 are scanned using confocal laser scanning microscopy (CLSM). For every point in the image, the nucleus density of BrdU-labeled nuclei and the total nucleus density of the neighborhood of that point are calculated from the BrdU and the To-Pro-3 signal, respectively. The ratio of these densities gives an indication of the amount of cell proliferation at that point. The automated measure is validated by comparing it with the ratio of BrdU-stained nuclei to the total number of nuclei obtained from a manual count.


RESULTS
A positive correlation is found between the automated measure and the ratios calculated from the manual counting (r = 0.86, P < 0.001). Calculating the topographical cell proliferation using the automated method is faster and does not suffer from interobserver variability.


CONCLUSIONS
Automated topographical cell proliferation analysis is a fast method to objectively find differences in cell proliferation within a tissue. This can be visualized by a topographical map that corresponds to the tissue under study.
van Raaij
E J
EJ
Hubrecht Laboratory, Netherlands Institute for Developmental Biology, Utrecht, The Netherlands.
ten Berge
D
D
Hage
W
W
Brouwer
A
A
Meijlink
F
F
Maintz
J B
JB
Verbeek
F J
FJ
D000818:Animals; D001973:Bromodeoxyuridine / Q000032:analysis / Q000378:metabolism; D002455:Cell Division; D002467:Cell Nucleus / Q000737:chemistry* / Q000378:metabolism; D004622:Embryo, Mammalian / Q000737:chemistry / Q000378:metabolism; D005260:Female; D019044:Image Cytometry / Q000379:methods*; D007091:Image Processing, Computer-Assisted; D008334:Mandible / Q000737:chemistry / Q000196:embryology / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D011247:Pregnancy; D015203:Reproducibility of Results
D001973:Bromodeoxyuridine