In situ nick translation method reveals DNA strand scission in HeLa cells following heat treatment.
DNA strand break in HeLa cells induced by heat was detected using the in situ nick translation method. The cells were incubated at 43 degrees C for various times (15, 30, 45, 60, 90 or 120 min) in Lab-Tek tissue culture chamber/slides and were fixed with ethanol/acetic acid on the slide glass. The break sites in DNA were translated artificially in the presence of Escherichia coli DNA polymerase I and 3H-labeled dTTP. The level of break sites in the DNA was visualized by autoradiographic observation of the grains. The DNA strand break appeared as early as 15 min, increased to 10.3-fold at 45 min of 43 degrees C treatment and this level related reciprocally to clonogenicity of the cell. The nick translation method thus provides a rapid in situ assay for determining heat-induced DNA damage of cultured cells, in a semi-quantitative manner.
Anai
H
H
Cancer Center of Kyushu University Hospital, Faculty of Medicine, Fukuoka, Japan.
Maehara
Y
Y
Sugimachi
K
K
D001426:Bacterial Proteins / Q000378:metabolism; D004249:DNA Damage; D004256:DNA Polymerase I / Q000378:metabolism; D004261:DNA Replication; D004273:DNA, Neoplasm / Q000032:analysis; D006367:HeLa Cells / Q000032:analysis; D006358:Hot Temperature; D006801:Humans; D006979:Hyperthermia, Induced
D001426:Bacterial Proteins; D004273:DNA, Neoplasm; D004256:DNA Polymerase I