Analysis of single-cell gel electrophoresis using laser-scanning microscopy.
The single-cell gel (SCG) assay has been shown to be a valuable technique to measure DNA-strand breaks in individual cells. Imaging of the assay is enhanced by laser-scanning microscopy (LSM) technology. Cells embedded in low melt point agarose were treated with 12.5 mM H2O2 to induce DNA-strand breaks. Following cell lysis and electrophoresis under alkaline conditions to allow single-stranded break detection, analysis of the resulting comets can provide an accurate method of comparing DNA-damage levels. The migration patterns of stained DNA were quantitated by LSM analysis. A statistically significant time-dependent dose-response relationship was clearly observed. LSM analysis of the SCG technique allows rapid, sensitive, and reproducible quantitation of single-stranded breaks and alkali-labile sites in the DNA of single cells.
O'Neill
K L
KL
Department of Microbiology, Brigham Young University, Provo UT 84602.
Fairbairn
D W
DW
Standing
M D
MD
D004249:DNA Damage; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D006367:HeLa Cells; D006801:Humans; D007834:Lasers; D008853:Microscopy / Q000379:methods*