5-Bromo-2′-deoxy-uridine Labeling and Detection Kit II Protocol & Troubleshooting

Product No. 11299964001

Protocol

Storage of samples after fixation

Storage of samples after fixation with ethanol fixative is possible for up to 2 weeks at +2 to +8 °C.

Troubleshooting

Reduction of high background

Using BrdU Labeling and Detection Kit II note the following:

a) Anti BrdU-POD will bind nonspecifically to some plastic material. Use sterile flat bottomed microplates that are anti-adsorbant (e.g., Maxisorp from NUNC or Costar). Always use flat bottomed sterile microplates. You can do a further blocking step with PBS/1% BSA before adding the antibody. Non-specific binding of BrdU to microplates can also be overcome by using poly-lysine to change the surface of the wells.

b) Another possible reason for high background: The anti-BrdU is not washed away by the washing steps. In this case, it may help to increase the number of washing steps (1-3- min, 4-5 times).

In addition to these points: Some cell lines show a nonspecific binding of the antibody at a cell concentration of more than 2x104 per ml. This may be due to the extracellular matrix. Reducing the cell number will in this case help to reduce background levels.

Materials

     
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