Ethidium Bromide (EtBr) revelation

Goal

Protocol

1. When the gel electrophoresis is finished, turn off the generator and put the gel in a plastic box.


2. Put the box in the EtBr area.



SAFETY AND USEFUL RECOMMANDATION
This box must not touch something contaminated with EtBr, because it could be used after for another electrophoresis and thus it would be put outside the EtBr area. However this box must be clean with water and alcohol before being reused.



3. Wear nitrile gloves over latex gloves, also wear the full facial protection.



SAFETY AND USEFUL RECOMMANDATION

Latex gloves do not protect hands from ethidium bromide, because it goes through the latex easily. That is why nitrile gloves are available in the laboratory. These gloves are more protective. However they are not completely protective. Thus it is recommended to wear them over the latex one. It is also recommended to prevent to have direct contact between gloves and product. The full facial protection prevents both possible spreding of solution and UV radiance.



4. Open the box and put the gel (with the spatula) in another box where there will be the EtBr solution.



SAFETY AND USEFUL RECOMMANDATION

The gel should be put in the box before the solution in order to prevent the formation of gaz and droplets. The spot, where this is done, is potentially contaminated with ethidium bromide. Each tool that would be used is in this place. In case of a lack of something it is important to think about its utilization. If it is necessary to use it, then it must not be used outside the ethidium bromide area, border with white and red lines. In the other way prevent the use of material taking from the outside of the area, because it would increase the risk of propagation of the ethidium bromide in the laboratory.



5. Prepare the EtBr solution (25 µL in 50 mL of water).



SAFETY AND USEFUL RECOMMANDATION

The ethidium bromide is stored in a concentrated way. This storage is done directly near the place the manipulation are done. Even if in the laboratory the safety of the storage should be improved, the product should all the time be stored at the same place which is indicated. Those manipulations are made inside the extracting hood. In case of incident like a spillage, users should not panic. The first thing is to prevent the spread of the liquid. The second is to conserve the liquid inside the extracting hood. The ethidium bromide is not a volatile compound. Thus there is little risk of inhalation. That is why if there is a spillage outside the extracting hood what should be done is exactly the same as what should be done inside. In case of spillage: Preventing the spread by using paper towel as dams around the pool. Absorbing the liquid with paper towel. Putting everything in the ethidium bromide garbage. Cleaning the spot with water and dry the place after. Cleaning twice with alcohol and putting everything in the ethidium bromide garbage. After having use the concentrated ethidium bromide it is very important to close properly the bottle and take it vertically.



6. After 10 min, put the gel under the UV lamp.



SAFETY AND USEFUL RECOMMANDATION

The gel is soaked with ethidium bromide. Consequently before taking it out, it is recommended to dry it a bit by waiting before putting it on the UV lamp. On the UV lamp there is also a window that has to be open before taking out the gel. In case of falling of the gel during its manipulation, two things are possible. If the gel can be used after, take the gel and put it on the UV lamp. Then clean the spot in the same way as explaining before. If the gel cannot be used, the entire gel must be clean before starting a new one. It is also possible that liquid could be spread on clothes. If it is the case it is recommended to throw the clothes in the ethidium bromide garbage.



7. Take a photo.



SAFETY AND USEFUL RECOMMANDATION

The UV lamp will be switched on. That is why people who do not have any UV protection must not be near the UV lamp. The person working on the computer must have the same individual protection as the manipulator, that is to say: blouse, nitrile gloves and full facial protection. What is also important is to have sleeves well over gloves and to have its blouse close to the neck. Not parts of the body must be directly exposed to the radiance. It is also recommended to close the window of the UV lamp and of the extracting hood.