A chemical method for fast and sensitive detection of DNA synthesis in vivo.
We have developed a method to detect DNA synthesis in proliferating cells, based on the incorporation of 5-ethynyl-2'-deoxyuridine (EdU) and its subsequent detection by a fluorescent azide through a Cu(I)-catalyzed [3 + 2] cycloaddition reaction ("click" chemistry). Detection of the EdU label is highly sensitive and can be accomplished in minutes. The small size of the fluorescent azides used for detection results in a high degree of specimen penetration, allowing the staining of whole-mount preparations of large tissue and organ explants. In contrast to BrdU, the method does not require sample fixation or DNA denaturation and permits good structural preservation. We demonstrate the use of the method in cultured cells and in the intestine and brain of whole animals.
Salic
Adrian
A
Department of Cell Biology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA. asalic@hms.harvard.edu
Mitchison
Timothy J
TJ
D000818:Animals; D004247:DNA / Q000032:analysis* / Q000096:biosynthesis* / Q000737:chemistry; D004261:DNA Replication; D003857:Deoxyuridine / Q000031:analogs & derivatives / Q000032:analysis / Q000737:chemistry; D051379:Mice; D015394:Molecular Structure; D041681:NIH 3T3 Cells; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013997:Time Factors
D004247:DNA; C031086:5-ethynyl-2'-deoxyuridine; D003857:Deoxyuridine