Safe Disposal of Ethidium Bromide 

Personal Protection 

When handling EtBr, always wear a lab coat, nitrile gloves, and chemical splash goggles. Proper skin and eye protection are needed when an ultraviolet (UV) light source is used. Avoid exposing unprotected skin and eyes to intense UV sources. Wear a face shield if UV source is pointing upwards. When working with a UV source for a long time, wrap up lab coat sleeves with tape or other means where the wrist could be exposed. 

Electrophoresis Gels 

Ethidium bromide in electrophoresis gels should be placed in an appropriate container for hazardous waste disposal. Please submit an online hazardous waste request via SciShield (formerly BioRAFT) once the waste is ready for a pickup.  

Aqueous Solutions 

Aqueous solutions can be decontaminated or deactivated using the following methods: 

Charcoal Filtration 
Filtering the aqueous ethidium bromide waste (free of other contaminants) through a bed of activated charcoal is a relatively simple and effective method for removal of ethidium bromide. The filtrate may be poured down the drain.  

Funnel Kit 
Commercial filter funnel kits use a graduated charcoal disk to track the amount of aqueous solution that ran through them. This is particularly useful for labs that generate large amounts of solutions at a time. The kit is available through Sigma-Aldrich or Thomas Scientific.  

Instructions for the funnel kit: 

1. Filter the ethidium bromide solution through the charcoal filter 
2. Pour filtrate down the drain 
3. Place charcoal filter in a sealed container and treat it as a hazardous waste 

Greenbag Kit 

Another simple charcoal filtration method is the Green Bag, manufactured by MP Biomedicals. The Greenbag Disposal Kit allows the rapid accumulation of ethidium bromide from large volumes of solutions into a small “tea bag”.   The “tea bag” contains activated carbon, which is then disposed of along with other solid hazardous wastes.  One kit includes 50 “teabags” which, in total, has the capacity to remove 500 mg of ethidium bromide from a solution. 

Instructions for the Greenbag kit:  

1. Place the Green Bag into the ethidium bromide solution 
2. Allow it to sit for the allotted time 
3. Pour filtrate down the drain 
4. Dispose of the Green Bag in a sealed container (mayonnaise jar) and label as a hazardous waste 

Chemical Deactivation 

Solutions containing ethidium bromide can be deactivated and rendered nonhazardous using one of the three methods. Deactivation may be confirmed by using UV light to detect fluorescence. The three methods are as follows: 

Armour Method 
The Armour Method is the simplest, but it is the most controversial due to one study showing that this method leaves traces of  mutagenic reactions (Lunn, G.and E. Sansone, Analytical Biochemistry, vol. 162, pp. 453-458, 1987). 

Armour method instructions:  

• Combine equal amounts of ethidium bromide solution and household bleach 
• Stir constantly for four hours or let sit for 2-3 days 
• Adjust pH to 6-10 with sodium hydroxide 
• Pour down drain with copious amounts of water.  (Because there is risk involved, we would like you to treat it as liquid Hazardous Waste.  Therefore, do not pour it down the drain!)  

Lunn and Sansone Method 

This method mainly uses hypophosphorus acid and sodium nitrile to remove the Ethidium Bromide.   

For each 100 ml of ethidium bromide solution: 

1. Add 5% hypophosphorus acid 
2. Add 12 ml of 0.5 M sodium nitrite 
3. Stir briefly and let stand for 20 hours 
4. Adjust pH to 6-10 using sodium hydroxide 
5. Pour down drain with copious amounts of water 

Crystals and Powders 

Label containers that contain crystals and powders as hazardous waste. Then, please submit an online hazardous waste request form once the waste is ready for a pickup. 

Contaminated Debris 

Contaminated debris includes gloves, bench paper and other non-labware items. Place contaminated debris into appropriate containers for hazardous waste disposal. All containers used for collection must have a top or cover that can be secured. Hazardous waste labels must be completed and affixed ontp the container.  Please submit an online hazardous waste request form once the waste is ready for pickup.  

Contaminated labware includes needles, pipettes, test tubes, etc. that are contaminated with ethidium bromide. Depending on the waste you generate, follow the procedures below: 

• Needles, scalpels, Pasteur pipettes, and other sharps contaminated with ethidium bromide should be disposed of directly into a sharps container. These will be disposed of like other sharps waste. 
• Volumetric or transfer pipettes, and other disposable glassware incidentally contaminated with ethidium bromide should be disposed of in a sharps container. Grossly contaminated (visibly contaminated) glassware should be washed with bleach before disposal in a sharps container. Rinsate containing <10 µg/ml of ethidium bromide may be poured down the drain to the sanitary sewer. Test tubes and centrifuge tubes contaminated with ethidium bromide should first be emptied with the liquid disposed of with the procedures given above or placed directly into a mayo jar. Empty incidentally contaminated tubes can be disposed of in the trash. Grossly contaminated (visibly contaminated) tubes should be collected with other laboratory debris, labeled, and disposed of as a hazardous waste. 
• Most of the other disposable labware (e.g. sample vials, disposable beakers, etc.) incidentally contaminated with Ethidium bromide may be disposed of in the normal trash. Grossly contaminated (visibly contaminated) disposable labware should be labeled and managed as a hazardous waste. 

IF THERE IS AN INCIDENT 

 Below are procedures to follow in case of contact: 

• In the case of eye contact, flush eye(s) with water for at least 15 minutes lifting upper and lower eyelids occasionally 
• If skin is exposed, remove contaminated clothing and wash skin with soap and water immediately 
• If Ethidium bromide vapors are inhaled, move to fresh air 
• If swallowed, get medical attention immediately 

Spill Clean-up

In case of a small spill, absorb freestanding liquid, use ultraviolet light to locate the spill, and prepare a decontamination solution by mixing 4.2 grams of sodium nitrite and 20 mL of hypophosphorous acid (50%) in 300 mL of water. To decontaminate the spill area, wash it with a soaked paper towel in the decontamination solution. Wash the spill area with wet paper towels (soaked in the decontamination solution) an additional 5 times, using fresh paper towels each time.  After this process, put all the decontamination solution in a bottle, label it as a hazardous waste and submit an online hazardous waste request form. 

 In case of a large spill, notify all people nearby and evacuate the laboratory immediately. Contact NUPD for assistance with clean-up. Post warning signs on all laboratory doors to notify others of the spill. Prevent unnecessary entry into the room until OARS personnel arrive. Explain the details of the incident to OARS personnel.  

Contaminated Equipment 

Laboratory equipment (e.g. transilluminators, laboratory floors and countertops, etc.) contaminated with an aqueous solution of ethidium bromide should be decontaminated using the spill clean-up procedures listed above. 

Waste Minimization 

The U.S. Congress has made waste minimization a national policy and goal of each waste generator. You as a generator of EtBr have the responsibility to minimize the waste you generate. Waste minimization has benefits such as decreasing your exposure to hazardous substances, protection of the environment, and decreasing the cost of disposal which can exceed the initial cost of the material by many times. 

 EtBr waste minimization is a priority. The following are suggestions for minimizing waste: 

• Substituting less or non-hazardous materials for hazardous materials. 
• Using dilute rather than concentrated solutions. 
• Using micro or semi-micro techniques. 
• Using computer modeling/simulation, films, videotapes or demonstrations instead of performing individual experiments. 
• Use commercially available filtrating vacuums or EtBr extractors to remove EtBr from buffers and other dilute aqueous solutions.