ETHIDIUM BROMIDE

Use of ethidium bromide for visualization of nucleic acids.

Potent mutagen. Moderately toxic.

Wear chemical safety goggles when using ethidium bromide. Avoid skin contact, ethidium
bromide may be absorbed through the skin. Use NATURAL rubber gloves.

Use only inside a chemical fume hood with adequate ventilation. Safety shower and eye wash should be easily accessible where ethidium bromide is used.

Store in a cool, dry place away from strong oxidizing agents. Keep containers tightly
closed. Use with adequate ventilation.

In case of contact with eyes, immediately flush eyes with copious amounts of water for
at least 15 minutes. In case of contact with skin, immediately wash skin with soap and
copious amounts of water. If inhaled, remove to fresh air. Call the OSU EHS-292 1284. 
If spill is on equipment, use ultraviolet light to locate spill, then use the decontamination 
procedure outlined below.

Wear protective clothing. 

Small spill: If in solution, absorb freestanding liquid. Use ultraviolet light to locate spill,
then use the decontamination procedure outlined below.

Large Spill: Notify others in room of spill. Evacuate room/immediate area. Call EHS for
cleanup. Post room with warning notifying others of spill. Prevent unnecessary entry into
area until arrival of EHS response team. Provide assistance and information to spill
responders.

Ethidium bromide is commonly used in molecular biology laboratories.  While it is not regulated as hazardous waste, the mutagenic properties of this substance may present a hazard if it is poured down the drain untreated or placed in the trash. 

Based on these considerations,  the following disposal procedures for ethidium bromide are recommended:

Electrophoresis Gels

Trace amounts of ethidium bromide in gels should not pose a hazard.  Higher concentrations, e.g., when the color of the gel is dark pink or red, should not be placed in laboratory trash.  EHS recommends that if the concentration is less than 0.1% ethidium bromide, gels may be placed in laboratory trash.  If concentration is equal to or greater than 0.1%, gels must be placed in Biohazard Burn Box for incineration.

     Less than 0.1% ethidium bromide:  place in laboratory trash 
     More than or equal to 0.1%:  place in biohazard box for incineration. 

Ethidium Bromide Solutions

Aqueous solutions containing <10ug/ml ethidium bromide can be released to the drain. 
Aqueous solutions containing >10ug/ml ethidium bromide should be filtered /deactivated using one of these methods: 

Charcoal Filtration

Filtering the aqueous ethidium bromide waste solutions, 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. 
There are two simple kits available for charcoal filtration: 

Funnel Kit 

Schleicher and Schuell supply a commercial  filter funnel kit that uses a packaged charcoal disk that is graduated for easily tracking the amount of aqueous solution calculated for a fixed quantities of ethidium bromide residue.  This is particularly useful for labs that generate large amounts of solutions at a time.  Kit is available through Schleicher and Schuell or VWR. 

• Filter the ethidium bromide solution through the charcoal filter.
• Pour filtrate down the drain.
• Place charcoal filter in a sealed bag (e.g., zip-lock) and place in biohazardous waste box for incineration.

"The Green Bag®" 

Another simple charcoal filtration method is the Green Bag, manufactured by BIO 101.  The Green Bag® Kit allows rapid and trouble-free concentration of ethidium bromide from large volumes of solutions into a small "tea" bag containing activated carbon which is then conveniently disposed along with other solid hazardous wastes. One kit has the capacity to remove 500 mg of ethidium bromide from solutions (10mg EtBr/bag).

• Place the Green Bag into the ethidium bromide solution.
• Allow to sit for the allotted time.
• Pour filtrate down the drain.
• Dispose of the Green back in the biohazardous waste box for incineration.

Chemical Neutralization

Armour Method

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 4-9 with sodium hydroxide.

Pour down drain with copious amounts of water.

Lunn and Sansone Method

Add 5% hypophosphorus acid.

Add 12 ml of 0.5 M sodium nitrate.

Stir briefly and let stand for 20 hours.

Adjust pH to 4-9 using sodium hydroxide.

Pour down drain with copious amounts of water.

Solutions containing heavy metals, organics, cyanides or sulfides should be disposed as hazardous waste. 

Gloves, Equipment and Debris

Contact OSU EHS-292-1284 for further disposal instructions.

PI approval before initial use of this material.

Wash the area with a paper towel soaked in a decontamination solution consisting of 4.2g
of sodium nitrite and 20 ml of hypophosphorous acid (50%) in 300 ml of water. Then wash
five times with wet paper towels using a fresh towel each time. Soak all the towels in
decontamination solution for 1 hour. Using a U- V light, check for completeness of
decontamination, and if satisfied, then call EHS for disposal of the decontamination
solution.

The decontamination solution must be prepared just prior to use.

If the decontamination solution (pH 1.8) is felt to be too corrosive for the surface to be
decontaminated, then use six rather than five washes following the decontamination
solution wipe. Again, soak all the towels in decontamination solution for at least 1 hour
before disposal.

Recommended.

Whenever ethidium bromide is mixed and stored outside its original container, it must be clearly marked
with its concentration and a description of its hazards. 

Use this PDF   and Avery Label # 5165 to print suitable label. 

Howard Hughes Medical Institute Lab Chemical Safety Summary-Ethidium Bromide

Printed MSDS-Ethidium Bromide

Princeton University EH&S