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Miniaturizing the comet assay with 3D vertical comets

Philippe Baert and Patric Van Oostveldt UGent (2003) CYTOMETRY PART A. 51A(1). p.26-34
abstract
Background: The comet or single-cell gel electrophoresis assay is a sensitive method for the detection of DNA damage. The main drawback of comet sampling is the low cell density necessary to prevent nucleus overlap after electrophoresis, which limits large-scale high throughput screening. Another problem may be inconsistent comet focusing. We investigated whether an approach based on three-dimensional (3D) confocal microscopy might be beneficial for these concerns. Methods: A vertical comet assay enabling three-dimensional confocal comet imaging of nuclei seeded at very high density was developed together with dedicated software algorithms to retrieve quantitative data at the single cell level. Results: Three-dimensional confocal comet imaging greatly relieved the user interactions of our nonautomated two-dimensional comet sampling procedure. Batches of comets were blindly sampled, and confocal sectioning improved the clarity of the images and the accuracy of comet sampling. A 1-Gy dose response was readily established. The sampling speed was competitive with that of commercial packages. Conclusions: Vertical comet imaging is a new concept for fast and user-friendly comet sampling that allows miniaturization of the assay. it may become an essential step toward high throughput screening and exploit the benefits of confocal imaging.
Please use this url to cite or link to this publication:
author
organization
year
type
journalArticle (original)
publication status
published
subject
keyword
FLUORESCENCE MICROSCOPY, ELECTROPHORESIS ASSAY, DNA-DAMAGE, single cell gel electrophoresis, confocal laser scanning microscopy, DNA damage, INDIVIDUAL CELLS, SLIDES, REPAIR, MOMENT
journal title
CYTOMETRY PART A
Cytom. Part A
volume
51A
issue
1
pages
26 - 34
Web of Science type
Article
Web of Science id
000182425600004
JCR category
BIOCHEMICAL RESEARCH METHODS
JCR impact factor
2.095 (2003)
JCR rank
22/49 (2003)
JCR quartile
2 (2003)
ISSN
0196-4763
DOI
10.1002/cyto.a.10006
language
English
UGent publication?
yes
classification
A1
copyright statement
I have transferred the copyright for this publication to the publisher
id
209019
handle
http://hdl.handle.net/1854/LU-209019
date created
2008-01-11 10:41:00
date last changed
2010-09-09 16:58:21
@article{209019,
  abstract     = {Background: The comet or single-cell gel electrophoresis assay is a sensitive method for the detection of DNA damage. The main drawback of comet sampling is the low cell density necessary to prevent nucleus overlap after electrophoresis, which limits large-scale high throughput screening. Another problem may be inconsistent comet focusing. We investigated whether an approach based on three-dimensional (3D) confocal microscopy might be beneficial for these concerns.
Methods: A vertical comet assay enabling three-dimensional confocal comet imaging of nuclei seeded at very high density was developed together with dedicated software algorithms to retrieve quantitative data at the single cell level.
Results: Three-dimensional confocal comet imaging greatly relieved the user interactions of our nonautomated two-dimensional comet sampling procedure. Batches of comets were blindly sampled, and confocal sectioning improved the clarity of the images and the accuracy of comet sampling. A 1-Gy dose response was readily established. The sampling speed was competitive with that of commercial packages.
Conclusions: Vertical comet imaging is a new concept for fast and user-friendly comet sampling that allows miniaturization of the assay. it may become an essential step toward high throughput screening and exploit the benefits of confocal imaging.},
  author       = {Baert, Philippe and Van Oostveldt, Patric},
  issn         = {0196-4763},
  journal      = {CYTOMETRY PART A},
  keyword      = {FLUORESCENCE MICROSCOPY,ELECTROPHORESIS ASSAY,DNA-DAMAGE,single cell gel electrophoresis,confocal laser scanning microscopy,DNA damage,INDIVIDUAL CELLS,SLIDES,REPAIR,MOMENT},
  language     = {eng},
  number       = {1},
  pages        = {26--34},
  title        = {Miniaturizing the comet assay with 3D vertical comets},
  url          = {http://dx.doi.org/10.1002/cyto.a.10006},
  volume       = {51A},
  year         = {2003},
}

Chicago
Baert, Philippe, and Patric Van Oostveldt. 2003. “Miniaturizing the Comet Assay with 3D Vertical Comets.” Cytometry Part A 51A (1): 26–34.
APA
Baert, Philippe, & Van Oostveldt, P. (2003). Miniaturizing the comet assay with 3D vertical comets. CYTOMETRY PART A, 51A(1), 26–34.
Vancouver
1.
Baert P, Van Oostveldt P. Miniaturizing the comet assay with 3D vertical comets. CYTOMETRY PART A. 2003;51A(1):26–34.
MLA
Baert, Philippe, and Patric Van Oostveldt. “Miniaturizing the Comet Assay with 3D Vertical Comets.” CYTOMETRY PART A 51A.1 (2003): 26–34. Print.