Product Catalog

The kit is used for the detection of BrdU incorporated into cellular DNA by immunocyto/histochemistry.

• Safe: No radioisotopes are used
• Easy to perform: Follows a standard immunohistochemistry protocol
• Sensitive: Denaturation of DNA with nucleases allows for highly sensitive detection of BrdU
• Flexible: Allows double-labeling protocols

Sample material: Cell culture: adherent cells, suspension cells, organ or explant cultures. Frozen or paraffin-embedded tissue sections (after in vivo labeling).
Specificity: Anti-BrdU monoclonal antibody specifically binds to 5-bromo-2'-deoxy-uridine, and shows crossreactivity with 5-iodo-2'-deoxy-uridine (10%). Anti-BrdU shows no crossreactivity with 5'-fluoro-2'-deoxy-uridine or any endogenous cellular component, such as thymidine or uridine.

Cell proliferation may be studied by monitoring the incorporation of a radioisotope, [ ³ H]-thymidine, into cellular DNA, followed by autoradiography.
Alternatively, 5-bromo-2'-deoxy-uridine (BrdU) may be used instead of thymidine. Cells that have incorporated BrdU into DNA are easily detected using a monoclonal antibody against BrdU and an enzyme- or fluorochrome-conjugated second antibody.

1. BrdU Labeling Reagent, 1,000x conc.
2. Washing Buffer concentrate, 10x
3. Incubation Buffer
4. Anti-BrdU, containing nucleases for DNA denaturation
5. Anti-mouse Ig-alkaline Phosphatase
6. NBT
7. BCIP

Samples prelabeled with BrdU are fixed with ethanol, then incubated with a monoclonal antibody to BrdU, which contains an optimized mixture of nucleases. These nucleases generate single-stranded DNA fragments that allow binding of the antibody to BrdU. Next, an alkaline phosphatase (AP)-labeled antibody to mouse immunoglobulin is added, then bound to the anti-BrdU antibody. The sample is then incubated with the AP substrate and NBT/ BCIP, which is metabolized to form a colored reaction product. The sample is evaluated using a phase-contrast microscope.

Figure 1: Principle of the BrdU Labeling and Detection Kit II (AP).