I am getting high DAB background staining. How can I decrease this background in the future?
High DAB background can be decreased by taking the following steps:
1) Make sure that the stringent wash is performed properly.
2) After hybridization, rinse the slides briefly in room temperature SSC buffer, then immerse slides for 5 minutes in SSC at 75°C.
3) Increase the temperature by 1°C per slide for more than 2 slides, but do not exceed 80°C.
4) For any other wash steps , 1X PBS with Tween 20 (0.025%) is recommended. Washing with any other reagents (1X PBS without Tween 20 or distilled water) can lead to unwanted background.
1) Make sure that the stringent wash is performed properly.
2) After hybridization, rinse the slides briefly in room temperature SSC buffer, then immerse slides for 5 minutes in SSC at 75°C.
3) Increase the temperature by 1°C per slide for more than 2 slides, but do not exceed 80°C.
4) For any other wash steps , 1X PBS with Tween 20 (0.025%) is recommended. Washing with any other reagents (1X PBS without Tween 20 or distilled water) can lead to unwanted background.