Waste

Items that go in biohazard waste

• Petri dishes containing media containing microbes and or plants
• Paper towels or gloves that have contacted microbes
• Pipet tips
• Plastic serological pipets

* Note: Any petri dishes containing hazardous substances or that have been stained with DAB must go in the mixed chemical biohazard waste stored by the chemical hood in room 369. See SOP for mixed chemical biohazards for a full list.

Before disposing of petri dishes containing bacteria or plants

• If you have one or two plates, tape the lids close.
• If there are several dishes, tape the whole stack together or put them in an empty petri dish sleeve and tape the sleeve closed.

If petri dishes contain Fungi

• Dishes must be closed with parafilm
• Or fill and empty petri dish sleeve with used dishes and tape the sleeve closed before putting into

 

When biohazard is half full or just past half it must be autoclaved

Using the autoclave:

• • Put an autoclave bin on a cart and open a new autoclave bag so that it is sitting in the bin.
  • Get a piece of masking tape, approx. 8 inches long. Take the lid off the biohazard waste and twist the top of the autoclave bag closed, but not tightly. There should be enough of gap to allow air to escape. Wrap the tape around the twist to keep the bag closed.
  • Pull the bag out and set it in the new bag on top of the autoclave bin. The second bag stays loose at the top. All waste must be double bagged before going into the autoclave.
  • Put a new bag in the biohazard waste.

 

• Autoclaves are located in rooms: 190, 213, 317, and 390

 

• If the autoclave is in use, look at the cycle that is currently running and set your timer so you know when it should complete. If they cycle has ended and it is safe to open the door, you can pull the other person’s contents out unless you think it is media – look at the user sheet for the lab who’s cycle just finished and let them know their cycle has completed.
• Autoclaves that use DI water have an interruption in water supply every 12 hours. Always check the bottom panel and see if the orange light is on. If the light is on, press the black button below it and wait for 12 to 15 minutes for it to heat up.
• Carefully load your waste into the autoclave, take care to not catch your bag on the edge of the rack guides along the sides.
• Close the door and push against the hinged side of the door to make sure it is flat against the opening before turning the lock. You want the lock to be nice and snug but not so tight you won’t be able to open it.
• The cycles may be different on other autoclaves so it’s important to read them before selecting if you are at a different machine. You’ll want the cycle that sterilizes for 20 mins and dries for 15. Press the number of the cycle you want, and it will display the settings. Press it again to confirm and start that cycle. Set you timer for 45 minutes.
• Fill out user sheet with the date, time cycle started, cycle selected, your initials / JW, and the room number.
• When cycle is complete, the pressure will be zero. You will see on the screen to wait 10 minutes liquids are hot. You can at this point unlock the door and slowly open it. A lot of hot steam will escape so stand back from the door. After steam has escaped you can reach in with heat protective gloves on and pull the bin of cooked waste out.

If the bag is a bit too full, it will be easier to move the cart in front of the opening and slide the bin onto the cart.

Allow the waste to cool before throwing into the dumpster out back by the loading dock.

Any petri dish stained with DAB qualifies as mixed chemical waste.

Follow these steps to neutralize any applied microbes or plant material that has been stained.

• Put on gloves, lab coat and eye protection.
• Move plates to be stained into the chemical hood.
• Lay down base neutralizing pad (looks like a large paper towel) from the spill kit to catch any drips.

 

Make a 20 percent bleach solution at the sink and take it to the fume hood.

• Estimate about 4 to 5 milliliters of solution per dish plus 5 milliliters for over pouring and drips that may occur.
• Remember, add bleach to water when making solutions.

Sample calculation

• Want to make 30 ml of a 20 percent bleach solution

 

• 20 percent = 0.2

 

• 0.2(30 ml) = 6 ml of bleach

 

• 30 – 6 = 24 ml of water

 

So, you would add 24 ml of water to a small beaker then add 6 ml of bleach and mix before using.

 

Adding bleach to petri dishes

• Pour enough bleach solution into to the petri dish to cover the surface of the media or plants. Cover and let sit for 10 minutes.
• If you are neutralizing a large quantity of plates, being the 10–minute wait period after applying bleach to the last dish.
• After the incubation period, you should notice the plates are now much lighter in color or clear, depending on how much stain was applied.
• Carefully put the plates into the mixed chemical waste container.

Mixed waste container must be properly labeled, lined with a clear heavy weight plastic bag and lid that twist closed all provided by REHS. When container is nearly full please let the lab, manager know. Lab manager will schedule REHS to pick up the container and bring a new one.