Product Catalog
5-Bromo-2'-deoxy-uridine Labeling and Detection Kit III | |
| Catalog Number | Pack size | |
|---|---|---|
| 11444611001 | 1 kit, (for up to 1,000 tests) | Order and Price Information |
Application
Use this ELISA assay kit to determine the amount of BrdU incorporated into cellular DNA.
Note: The assay is performed in 96-well microplates.
Benefits
- Safer, since the kit does not use radioisotopes.
- Accurate, since results generated with this assay strongly correlate to those obtained with the [3 H]-thymidine method. (See Figure 2 below.)
- Sensitive. This assay and the [3 H]-thymidine assay are equally sensitive. (See Figure 2 below.)
- Fast, since results can be read with a multi-well ELISA reader, allowing simultaneous processsing of a large number of samples
- Easy, since the assay uses a standard cell ELISA protocol.
- Economical, since the assay requires no expensive equipment or additional reagents (e.g ., scintillation fluid).
Product Description
Sensitivity: The sensitivity of the BrdU Labeling and Detection Kit III is comparable to that of the traditional [3 H]-thymidine assay. (See Figure 2 below.)
Sample material: Adherent or suspension cells cultured in 96-well microplates.
Background Information
Proliferation in cell populations may be studied by incorporating the radioisotope [3H]-thymidine into cellular DNA. The amount of radioactive thymidine incorporated is determined by scintillation counting.
Alternatively , 5-bromo-2'-deoxy-uridine may be incorporated into cellular DNA. The amount of BrdU incorporated is determined by a standard ELISA protocol, which involves "tagging" the incorporated nucleotide with an anti-BrdU antibody .
Contents
- BrdU Labeling Reagent, 1000x
- Washing Buffer concentrate, 10x
- Incubation Buffer
- Nucleases
- Anti-BrdU-POD, Fab fragments
- Substrate Buffer
- ABTS Substrate
- Substrate Enhancer
Principle
Cells cultured in a 96-well microplate are incubated with BrdU (Figure 1).
The labeled cells are fixed with ethanol. Prior to incubation with a monoclonal
antibody to BrdU, DNA is partially digested with nucleases to allow the antibody
to access BrdU. Next, the anti-BrdU antibody [labeled with peroxidase (POD)] is
added. Finally, the POD substrate ABTS is added. POD catalyzes the cleavage of
ABTS, producing a colored reaction product. The absorbance of the samples (at
approx. 405 nm) is determined with a standard microplate (ELISA) reader.
Figure 1: Principle of the BrdU Labeling and Detection Kit III (POD).
Typical Experiment
Figure 2: Comparison of the sensitivity of the BrdU and [3H]-thymidine methods in measuring proliferation, as shown in hEGF (left figure) and mIL-6 (right figure) stimulation assays.
Left: Proliferation of AKR-2B cells (mouse fibroblast cell line) in response to recombinant human epidermal growth factor (hEGF). Proliferation was measured with either the BrdU Labeling and Detection Kit III (red) or the [3 H]-thymidine method (blue).
Right: Proliferation of 7TD1 cells (mouse-mouse hybridoma) in response to recombinant mouse interleukin-6 (mIL-6). Proliferation was measured with either the BrdU Labeling and Detection Kit III (red) or the [3 H]-thymidine method (blue).
Conclusion: In both cases, the cell proliferation assay with the nonradioactive BrdU kit was just as sensitive as the radioactive thymidine assay.
Additional Information
Special Interest Sites
Product Literature
Instructions for Use and Material Safety Data Sheets
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