Nuclear and nonnuclear incorporation of 5-bromo-2'-deoxyuridine in live cells.
Nuclear and nonnuclear incorporation of 5-bromo-2'-deoxyuridine in live cells. Bovine pulmonary arterial endothelial (BPAE) cells were labeled with 5-bromo-2'-deoxyuridine (BrdU, Cat. no. B23151) applied at a concentration of 10 µM for 30 minutes. After fixation with 4% formaldehyde in phosphate-buffered saline for 30 minutes, chromatin was denatured by treatment with 2 M HCl for 20 minutes. Incorporated BrdU was detected with mouse monoclonal anti-bromodeoxyuridine antibody (Cat. no. A21300) followed by HRP-conjugated goat anti–mouse IgG antibody and Oregon Green® 488 tyramide (TSA Kit #9, Cat. no. T20919). Tyramide labeling was further amplified and converted for visualization by bright-field microscopy by detection of the Oregon Green® 488 dye hapten using the HRP conjugate of anti-fluorescein/Oregon Green® antibody (Cat. no. A21253) and diaminobenzidine (DAB) staining. Both nuclear and non-nuclear (presumably mitochondrial) incorporation of BrdU is clearly visible in the resulting image.