BrdU Mouse anti-Chemical, APC, Clone: BU20A, eBioscience™

Description

Following the incubation, harvest the cells and wash once with 1X PBS.3. Stain surface molecules according to the Surface Staining Protocol.4. Wash in cold Flow Cytometry Staining Buffer or 1X PBS.5. Resuspend the cell pellet by pulse vortexing. Then add 1 mL of freshly prepared Foxp3 Fixation/Permeabilization Buffer (cat. 00-5521) to each sample. pulse vortex again.6. Incubate for 30 to 60 minutes at 2-8°C in the dark.7. Wash once with cold Flow Cytometry Staining Buffer followed by centrifugation. Decant the supernatant.8. Resuspend the cell pellet with 100 µL Flow Cytometry Staining Buffer containing 30 µg of Dnase I.9. Incubate for 1 hr at 37°C and then wash.10. Stain cells with anti-BrdU antibody for 30 min to 1 hr and then wash.10. Analyze the samples. Excitation: 633-647 nm; Emission: 660 nm; Laser: Red Laser. Filtration: 0.2 µm post-manufacturing filtered. Bromodeoxyuridine (5-bromo-2-deoxyuridine, BrdU) is a synthetic nucleoside that is an analogue of thymidine. BrdU is commonly used in the detection of proliferating cells in living tissues, and can be incorporated into the newly synthesized DNA of replicating cells (during the S phase of the cell cycle), substituting for thymidine during DNA replication. Antibodies specific for BrdU can then be used to detect the incorporated chemical, thus indicating cells that were actively replicating their DNA. Binding of the antibody requires denaturation of the DNA, usually by exposing the cells to acid or heat.